1-(2-phenoxymethylphenyl)piperazine compounds

ABSTRACT

The invention relates to compounds of formula I: 
                         
where a, and R 1  through R 6  are as defined in the specification, or a pharmaceutically acceptable salt thereof. The compounds of formula I are serotonin and norepinephrine reuptake inhibitors. The invention also relates to pharmaceutical compositions comprising such compounds; methods of using such compounds; and process and intermediates for preparing such compounds.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application No.61/293,949, filed on Jan. 11, 2010; the entire disclosure of which isincorporated herein by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to 1-(2-phenoxymethylphenyl)piperazinecompounds having activity as serotonin (5-HT) and norepinephrine (NE)reuptake inhibitors. The invention also relates to pharmaceuticalcompositions comprising such compounds, processes and intermediates forpreparing such compounds and methods of using such compounds to treat apain disorder, such as neuropathic pain, and other ailments.

2. State of the Art

Pain is an unpleasant sensory and emotional experience associated withactual or potential tissue damage, or described in terms of such damage(International Association for the Study of Pain (IASP), PainTerminology). Chronic pain persists beyond acute pain or beyond theexpected time for an injury to heal (American Pain Society. “PainControl in the Primary Care Setting.” 2006:15). Neuropathic pain is paininitiated or caused by a primary lesion or dysfunction in the nervoussystem. Peripheral neuropathic pain occurs when the lesion ordysfunction affects the peripheral nervous system and centralneuropathic pain when the lesion or dysfunction affects the centralnervous system (IASP).

Several types of therapeutic agents are currently used to treatneuropathic pain including, for example, tricyclic antidepressants(TCAs), serotonin and norepinephrine reuptake inhibitors (SNRIs),calcium channel ligands (e.g., gabapentin and pregabalin), topicallidocaine, and opioid agonists (e.g., morphine, oxycodone, methadone,levorphanol and tramadol). However, neuropathic pain can be verydifficult to treat with no more than 40-60% of patients achieving, atbest, partial relief of their pain (Dworkin et al. (2007) Pain132:237-251). Moreover, all of the therapeutic agents currently used totreat neuropathic pain have various side effects (e.g., nausea,sedation, dizziness and somnolence) that can limit their effectivenessin some patients (Dworkin et al. supra).

SNRIs, such as duloxetine and venlafaxine, are often used as first linetherapy for treating neuropathic pain. These agents inhibit the reuptakeof both serotonin (5-hydroxytrypamine, 5-HT) and norepinephrine (NE) bybinding to the serotonin and norepinephrine transporters (SERT and NET,respectively). However, both duloxetine and venlafaxine have higheraffinity for SERT relative to NET (Vaishnavi et al. (2004) Biol.Psychiatry 55(3):320-322).

Preclinical studies suggest that inhibition of both SERT and NET may benecessary for maximally effective treatment of neuropathic and otherchronic pain states (Jones et al. (2006) Neuropharmacology51(7-8):1172-1180; Vickers et al. (2008) Bioorg. Med. Chem. Lett.18:3230-3235; Fishbain et al. (2000) Pain Med. 1(4):310-316; andMochizucki (2004) Human Psychopharmacology 19:S15-S19). However, inclinical studies, the inhibition of SERT has been reported to be relatedto nausea and other side effects (Greist et al. (2004) Clin. Ther.26(9):1446-1455). Thus, therapeutic agents having more balanced SERT andNET affinity or slightly higher NET affinity are expected to beparticularly useful for treating chronic pain while producing fewer sideeffects, such as nausea.

Thus, a need exists for novel compounds that are useful for treatingchronic pain, such as neuropathic pain. In particular, a need exists fornovel compounds that are useful for treating chronic pain and that havereduced side effects, such as nausea. A need also exists for noveldual-acting compounds that inhibit both SERT and NET with fairly highaffinity (e.g., pK_(i)≧7.0 or K_(i)≦100 nM) and balanced inhibition(e.g., a SERT/NET binding K_(i) ratio of 0.1 to 100).

SUMMARY OF THE INVENTION

The present invention provides novel compounds that have been found topossess serotonin reuptake inhibitory activity and norepinephrinereuptake inhibitory activity. Accordingly, compounds of the inventionare expected to be useful and advantageous as therapeutic agents forthose diseases and disorders that can be treated by inhibition of theserotonin and/or norepinephrine transporter, such as neuropathic pain.

One aspect of the invention relates to compounds of formula I:

where: a is 0, 1, or 2; each R¹ is independently halo ortrifluoromethyl; R²⁻⁶ are independently hydrogen, halo, —C₁₋₆alkyl,—CF₃, —C₂₋₆alkynyl, —O—C₁₋₆alkyl, —OCF₃, —C₁₋₄alkylene-O—C₁₋₄alkyl,—C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl, —C₀₋₆alkylene-OH, —CN,—C₀₋₂alkylene-COOH, —CHO, —C(O)—C₁₋₆alkyl, —C(O)O—C₁₋₄alkyl, —CH₂SH,—S—C₁₋₆alkyl, —C₁₋₄alkylene-S—C₁₋₄alkyl, —SO₂—C₁₋₆alkyl, —SO₂NRR,—NHSO₂R, —C₀₋₁alkylene-NRR, —NHC(O)—C₁₋₆alkyl, —C(O)NRR, or —NO₂; andeach R is independently hydrogen or —C₁₋₄alkyl; or a pharmaceuticallyacceptable salt thereof; with the proviso that the compound of formula Iis not 1-(2-phenoxymethylphenyl)piperazine,1-[2-(3-fluorophenoxymethyl)phenyl]piperazine, or1-[2-(2,3-difluorophenoxymethyl)phenyl]-piperazine.

Yet another aspect of the invention relates to pharmaceuticalcompositions comprising a pharmaceutically acceptable carrier and acompound of the invention. Such compositions may optionally containother active agents such as anti-Alzheimer's agents, anticonvulsants,antidepressants, anti-Parkinson's agents, dual serotonin-norepinephrinereuptake inhibitors, non-steroidal anti-inflammatory agents,norepinephrine reuptake inhibitors, opioid agonists, opioid antagonists,selective serotonin reuptake inhibitors, sodium channel blockers,sympatholytics, and combinations thereof. Accordingly, in yet anotheraspect of the invention, a pharmaceutical composition comprises acompound of the invention, a second active agent, and a pharmaceuticallyacceptable carrier. Another aspect of the invention relates to acombination of active agents, comprising a compound of the invention anda second active agent. The compounds of the invention can be formulatedtogether or separately from the additional agent(s). When formulatedseparately, a pharmaceutically acceptable carrier may be included withthe additional agent(s). Thus, yet another aspect of the inventionrelates to a combination of pharmaceutical compositions, the combinationcomprising: a first pharmaceutical composition comprising a compound offormula I or a pharmaceutically acceptable salt thereof and a firstpharmaceutically acceptable carrier; and a second pharmaceuticalcomposition comprising a second active agent and a secondpharmaceutically acceptable carrier. The invention also relates to a kitcontaining such pharmaceutical compositions, for example where the firstand second pharmaceutical compositions are separate pharmaceuticalcompositions.

Compounds of the invention possess serotonin reuptake inhibitoryactivity and norepinephrine reuptake inhibitory activity, and aretherefore expected to be useful as therapeutic agents for treatingpatients suffering from a disease or disorder that is treated by theinhibition of the serotonin and/or the norepinephrine transporter. Thus,one aspect of the invention relates to a method of treating: a paindisorder such as neuropathic pain or fibromyalgia; a depressive disordersuch as major depression; an affective disorder such as an anxietydisorder; attention deficit hyperactivity disorder; a cognitive disordersuch as dementia; stress urinary incontinence; chronic fatigue syndrome;obesity; or vasomotor symptoms associated with menopause, comprisingadministering to a patient a therapeutically effective amount of acompound of the invention.

Still another aspect of the invention relates to a method for inhibitingserotonin reuptake in a mammal comprising administering to the mammal, aserotonin transporter-inhibiting amount of a compound of the invention.Yet another aspect of the invention relates to a method for inhibitingnorepinephrine reuptake in a mammal comprising administering to themammal, a norepinephrine transporter-inhibiting amount of a compound ofthe invention. And another aspect of the invention is directed to amethod for inhibiting serotonin reuptake and norepinephrine reuptake ina mammal comprising administering to the mammal, a serotonintransporter- and norepinephrine transporter-inhibiting amount of acompound of the invention.

Among the compounds of formula I, compounds of particular interest arethose having an inhibitory constant (pK_(i)) at SERT and at NET greaterthan or equal to about 5.0; and in another embodiment having a pK_(i) atSERT and at NET greater than or equal to about 7.0. In anotherembodiment, compounds of interest have balanced SERT and NET activity,i.e., have the same pK_(i) value at both SERT and NET±0.5. Furthercompounds of particular interest are those having a serotonin reuptakeinhibition pIC₅₀ values of greater than or equal to about 7.0 and anorepinephrine reuptake inhibition pIC₅₀ values of greater than or equalto about 7.0.

Since compounds of the invention possess serotonin reuptake inhibitoryactivity and norepinephrine reuptake inhibitory activity, such compoundsare also useful as research tools. Accordingly, one aspect of theinvention relates to methods of using the compounds of the invention asresearch tools, comprising conducting a biological assay using acompound of the invention. Compounds of the invention can also be usedto evaluate new chemical compounds. Thus another aspect of the inventionrelates to a method of evaluating a test compound in a biological assay,comprising: (a) conducting a biological assay with a test compound toprovide a first assay value; (b) conducting the biological assay with acompound of the invention to provide a second assay value; wherein step(a) is conducted either before, after or concurrently with step (b); and(c) comparing the first assay value from step (a) with the second assayvalue from step (b). Exemplary biological assays include a serotoninreuptake assay and a norepinephrine reuptake assay. Still another aspectof the invention relates to a method of studying a biological system orsample comprising serotonin transporters, norepinephrine transporters,or both, the method comprising: (a) contacting the biological system orsample with a compound of the invention; and (b) determining the effectscaused by the compound on the biological system or sample.

The invention also relates to processes and intermediates useful forpreparing compounds of the invention. Accordingly, one aspect of theinvention relates to a process for preparing compounds of formula I, theprocess comprising deprotecting a compound of formula IV:

or a salt thereof, to provide compounds of formula I, where a and R¹through R⁶ are as defined for formula I and P is an amino-protectinggroup selected from t-butoxycarbonyl, trityl, benzyloxycarbonyl,9-fluorenylmethoxycarbonyl, formyl, and benzyl. In other aspects, theinvention relates to novel intermediates used in such processes.

Yet another aspect of the invention relates to the use of compounds ofthe invention for the manufacture of medicaments, especially for themanufacture of medicaments useful for treating pain disorders,depressive disorders, affective disorders, attention deficithyperactivity disorder, cognitive disorders, stress urinaryincontinence, for inhibiting serotonin reuptake in a mammal, or forinhibiting norepinephrine reuptake in a mammal Still another aspect ofthe invention relates to the use of compounds of the invention asresearch tools. Other aspects and embodiments of the invention aredisclosed herein.

DETAILED DESCRIPTION OF THE INVENTION Definitions

When describing the compounds, compositions, methods and processes ofthe invention, the following terms have the following meanings unlessotherwise indicated. Additionally, as used herein, the singular forms“a,” “an” and “the” include the corresponding plural forms unless thecontext of use clearly dictates otherwise. The terms “comprising”,“including,” and “having” are intended to be inclusive and mean thatthere may be additional elements other than the listed elements. Allnumbers expressing quantities of ingredients, properties such asmolecular weight, reaction conditions, and so forth used herein are tobe understood as being modified in all instances by the term “about,”unless otherwise indicated. Accordingly, the numbers set forth hereinare approximations that may vary depending upon the desired propertiessought to be obtained by the present invention. At least, and not as anattempt to limit the application of the doctrine of equivalents to thescope of the claims, each number should at least be construed in lightof the reported significant digits and by applying ordinary roundingtechniques.

The term “alkyl” means a monovalent saturated hydrocarbon group whichmay be linear or branched. Unless otherwise defined, such alkyl groupstypically contain from 1 to 10 carbon atoms and include, for example,—C₁₋₂alkyl, —C₁₋₃alkyl, —C₁₋₄alkyl, —C₁₋₆alkyl, and —C₁₋₈alkyl.Representative alkyl groups include, by way of example, methyl, ethyl,n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl,n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl, and the like.

When a specific number of carbon atoms is intended for a particular termused herein, the number of carbon atoms is shown preceding the term assubscript. For example, the term “—C₁₋₆alkyl” means an alkyl grouphaving from 1 to 6 carbon atoms, and the term “—C₁₋₄alkylene” means analkylene group having from 1 to 4 carbon atoms, where the carbon atomsare in any acceptable configuration.

The term “alkylene” means a divalent saturated hydrocarbon group thatmay be linear or branched. Unless otherwise defined, such alkylenegroups typically contain from 0 to 10 carbon atoms and include, forexample, —C₀₋₁alkylene, —C₀₋₂alkylene, —C₀₋₃alkylene, —C₀₋₆alkylene,—C₁₋₄alkylene, —C₂₋₄alkylene and —C₁₋₆alkylene. Representative alkylenegroups include, by way of example, methylene (—CH₂—), ethylene(—CH₂—CH₂—), propylene (—CH₂—CH₂—CH₂—, —CH₂(CH₃)—CH₂—, or—CH₂—CH₂(CH₃)—), butylene (—CH₂—CH₂—CH₂—CH₂—, —CH₂(CH₃)—CH₂—CH₂—,—CH₂—CH₂(CH₃)—CH₂—, —CH₂—CH₂—CH₂(CH₃)—, —CH₂—CH₂(CH₂CH₃)—, or—CH₂(CH₂CH₃)—CH₂—), pentylene, and the like. It is understood that whenthe alkylene term includes zero carbons such as —C₀₋₁alkylene-,—C₀₋₃alkylene- or —C₀₋₆alkylene-, such terms are intended to include theabsence of carbon atoms, i.e., the alkylene group is not present exceptfor a covalent bond attaching the groups separated by the alkylene term.

The term “alkynyl” means a monovalent unsaturated hydrocarbon groupwhich may be linear or branched and which has at least one, andtypically 1, 2 or 3, carbon-carbon triple bonds. Unless otherwisedefined, such alkynyl groups typically contain from 2 to 10 carbon atomsand include, for example, —C₂₋₄alkynyl, —C₂₋₆alkynyl and —C₃₋₁₀alkynyl.Representative alkynyl groups include, by way of example, ethynyl,n-propynyl, i-butynyl, n-but-2-ynyl, n-hex-3-ynyl, and the like.

The term “halo” means fluoro, chloro, bromo and iodo.

As used herein, the phrase “of the formula”, “having the formula” or“having the structure” is not intended to be limiting and is used in thesame way that the term “comprising” is commonly used.

The term “pharmaceutically acceptable” refers to a material that is notbiologically or otherwise unacceptable when used in the invention. Forexample, the term “pharmaceutically acceptable carrier” refers to amaterial that can be incorporated into a composition and administered toa patient without causing unacceptable biological effects or interactingin an unacceptable manner with other components of the composition. Suchpharmaceutically acceptable materials typically have met the requiredstandards of toxicological and manufacturing testing, and include thosematerials identified as suitable inactive ingredients by the U.S. Foodand Drug Administration.

The term “pharmaceutically acceptable salt” means a salt prepared from abase or an acid which is acceptable for administration to a patient,such as a mammal (e.g., salts having acceptable mammalian safety for agiven dosage regime). However, it is understood that the salts coveredby the invention are not required to be pharmaceutically acceptablesalts, such as salts of intermediate compounds that are not intended foradministration to a patient. Pharmaceutically acceptable salts can bederived from pharmaceutically acceptable inorganic or organic bases andfrom pharmaceutically acceptable inorganic or organic acids. Inaddition, when a compound of formula I contains both a basic moiety,such as an amine, and an acidic moiety such as a carboxylic acid,zwitterions may be formed and are included within the term “salt” asused herein. Salts derived from pharmaceutically acceptable inorganicbases include ammonium, calcium, copper, ferric, ferrous, lithium,magnesium, manganic, manganous, potassium, sodium, and zinc salts, andthe like. Salts derived from pharmaceutically acceptable organic basesinclude salts of primary, secondary and tertiary amines, includingsubstituted amines, cyclic amines, naturally-occurring amines, and thelike, such as arginine, betaine, caffeine, choline,N,N-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol,2-dimethylaminoethanol, ethanolamine, ethylenediamine,N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine,hydrabamine, isopropylamine, lysine, methylglucamine, morpholine,piperazine, piperadine, polyamine resins, procaine, purines,theobromine, triethylamine, trimethylamine, tripropylamine,tromethamine, and the like. Salts derived from pharmaceuticallyacceptable inorganic acids include salts of boric, carbonic, hydrohalic(hydrobromic, hydrochloric, hydrofluoric or hydroiodic), nitric,phosphoric, sulfamic and sulfuric acids. Salts derived frompharmaceutically acceptable organic acids include salts of aliphatichydroxyl acids (e.g., citric, gluconic, glycolic, lactic, lactobionic,malic, and tartaric acids), aliphatic monocarboxylic acids (e.g.,acetic, butyric, formic, propionic and trifluoroacetic acids), aminoacids (e.g., aspartic and glutamic acids), aromatic carboxylic acids(e.g., benzoic, p-chlorobenzoic, diphenylacetic, gentisic, hippuric, andtriphenylacetic acids), aromatic hydroxyl acids (e.g., o-hydroxybenzoic,p-hydroxybenzoic, 1-hydroxynaphthalene-2-carboxylic and3-hydroxynaphthalene-2-carboxylic acids), ascorbic, dicarboxylic acids(e.g., fumaric, maleic, oxalic and succinic acids), glucuronic,mandelic, mucic, nicotinic, orotic, pamoic, pantothenic, sulfonic acids(e.g., benzenesulfonic, camphorsulfonic, edisylic, ethanesulfonic,isethionic, methanesulfonic, naphthalenesulfonic,naphthalene-1,5-disulfonic, naphthalene-2,6-disulfonic andp-toluenesulfonic acids), xinafoic acid, and the like.

The term “solvate” means a complex or aggregate formed by one or moremolecules of a solute, e.g., a compound of formula I or apharmaceutically acceptable salt thereof, and one or more molecules of asolvent. Such solvates are typically crystalline solids having asubstantially fixed molar ratio of solute and solvent. Representativesolvents include, by way of example, water, methanol, ethanol,isopropanol, acetic acid, and the like. When the solvent is water, thesolvate formed is a hydrate.

The term “therapeutically effective amount” means an amount sufficientto effect treatment when administered to a patient in need thereof,i.e., the amount of drug needed to obtain the desired therapeuticeffect. For example, a therapeutically effective amount for treatingneuropathic pain is an amount of compound needed to, for example,reduce, suppress, eliminate or prevent the symptoms of neuropathic painor to treat the underlying cause of neuropathic pain. On the other hand,the term “effective amount” means an amount sufficient to obtain adesired result, which may not necessary be a therapeutic result. Forexample, when studying a system comprising a norepinephrine transporter,an “effective amount” may be the amount needed to inhibit norepinephrinereuptake.

The term “treating” or “treatment” as used herein means the treating ortreatment of a disease or medical condition (such as neuropathic pain)in a patient, such as a mammal (particularly a human), that includes oneor more of the following: (a) preventing the disease or medicalcondition from occurring, i.e., prophylactic treatment of a patient; (b)ameliorating the disease or medical condition, i.e., eliminating orcausing regression of the disease or medical condition in a patient; (c)suppressing the disease or medical condition, i.e., slowing or arrestingthe development of the disease or medical condition in a patient; or (d)alleviating the symptoms of the disease or medical condition in apatient. For example, the term “treating neuropathic pain” would includepreventing neuropathic pain from occurring, ameliorating neuropathicpain, suppressing neuropathic pain, and alleviating the symptoms ofneuropathic pain. The term “patient” is intended to include thosemammals, such as humans, that are in need of treatment or diseaseprevention, that are presently being treated for disease prevention ortreatment of a specific disease or medical condition, as well as testsubjects in which compounds of the invention are being evaluated orbeing used in a assay, for example an animal model.

All other terms used herein are intended to have their ordinary meaningas understood by those of ordinary skill in the art to which theypertain.

In one aspect, this invention relates to compounds of formula I:

or a pharmaceutically acceptable salt thereof.

As used herein, the term “compound of the invention” or “compounds ofthe invention” include all compounds encompassed by formulas I, II, III,and IV, such as the species embodied in formulas Ia and all othersubspecies of such formula. In addition, when a compound of theinvention contain a basic or acidic group (e.g., amino or carboxylgroups), the compound can exist as a free base, free acid, a zwitterion,or in various salt forms. All such salt forms are included within thescope of the invention. Accordingly, those skilled in the art willrecognize that reference to compounds herein, for example, reference toa “compound of the invention” or a “compound of formula I” includes acompound of formula I as well as pharmaceutically acceptable salts ofthat compound unless otherwise indicated. Furthermore, solvates are alsoincluded within the scope of this invention.

Compounds of the invention, as well as those compounds used in theirsynthesis, may also include isotopically-labeled compounds, i.e., whereone or more atoms have been enriched with atoms having an atomic massdifferent from the atomic mass predominately found in nature. Examplesof isotopes that may be incorporated into the compounds of theinvention, for example, include, but are not limited to, ²H, ³H, ¹³C,¹⁴C, ¹⁵N, ¹⁸O, ¹⁷O, ³⁵S, ³⁶Cl, and ¹⁸F. Of particular interest arecompounds of formula I enriched in tritium or carbon-14 which can beused, for example, in tissue distribution studies; compounds of formulaI enriched in deuterium especially at a site of metabolism resulting,for example, in compounds having greater metabolic stability; andcompounds of formula I enriched in a positron emitting isotope, such as¹¹C, ¹⁸F, ¹⁵O ¹³N, which can be used, for example, in Positron EmissionTopography (PET) studies.

The compounds of the invention have been found to possess serotoninreuptake inhibitory activity and norepinephrine reuptake inhibitoryactivity. Among other properties, such compounds are expected to beuseful as therapeutic agents for treating chronic pain, such asneuropathic pain. By combining dual activity into a single compound,double therapy can be achieved, i.e., serotonin reuptake inhibitoryactivity and norepinephrine reuptake inhibitory activity, using a singleactive component. Since pharmaceutical compositions containing oneactive component are typically easier to formulate than compositionscontaining two active components, such single-component compositionsprovide a significant advantage over compositions containing two activecomponents.

Many combined serotonin and norepinephrine reuptake inhibitors (SNRIs)are more selective for SERT than for NET. For example, milnacipran,duloxetine, and venlafaxine exhibit 2.5-fold, 10-fold, and 100-foldselectivity (measured as pK_(i)) for SERT over NET, respectively. Some,however, are less selective, such as bicifadine, which has a pK_(i) atSERT of 7.0 and a pK_(i) at NET of 6.7. Since it may be desirable toavoid selective compounds, in one embodiment of the invention thecompounds have a more balanced SERT and NET activity, i.e., have thesame pK_(i) value at both SERT and NET±0.5.

The compounds described herein have typically been named using theAutoNom feature of the commercially-available MDL® ISIS/Draw software(Symyx, Santa Clara, Calif.). Typically, compounds of the invention havebeen named as 1-(2-phenoxymethylphenyl)piperazines.

Representative Embodiments

The following substituents and values are intended to providerepresentative examples of various aspects and embodiments of theinvention. These representative values are intended to further defineand illustrate such aspects and embodiments and are not intended toexclude other embodiments or to limit the scope of the invention. Inthis regard, the representation that a particular value or substituentis preferred is not intended in any way to exclude other values orsubstituents from the invention unless specifically indicated.

In one aspect, this invention relates to compounds of formula I:

The integer a can be zero, one or two. Thus, in one embodiment there areno R¹ groups present. In another embodiment, there is one R¹ groupselected from halo and trifluoromethyl; and in yet another embodimentthere are two R¹ groups independently selected from halo andtrifluoromethyl.

The R², R³, R⁴, R⁵, and R⁶ groups are independently hydrogen, halo,—C₁₋₆alkyl, —CF₃, —C₂₋₆alkynyl, —O—C₁₋₆alkyl, —OCF₃,—C₁₋₄alkylene-O—C₁₋₄alkyl, —C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl,—C₀₋₆alkylene-OH, —CN, —C₀₋₂alkylene-COOH, —CHO, —C(O)—C₁₋₆alkyl,—C(O)O—C₁₋₄alkyl, —CH₂SH, —S—C₁₋₆alkyl, —C₁₋₄alkylene-S—C₁₋₄alkyl,—SO₂—C₁₋₆alkyl, —SO₂NRR, —NHSO₂R, —C₀₋₁alkylene-NRR, —NHC(O)—C₁₋₆ alkyl,—C(O)NRR, or —NO₂. Each R group is independently hydrogen or —C₁₋₄alkyl.

In some embodiments of the invention, one or more positions on thephenyl ring are substituted with a non-hydrogen moiety. For example, onesuch embodiment may be described by stating that that “R² is anon-hydrogen moiety”. It is understood that this means that R² can beany of the non-hydrogen moieties defined in formula I, i.e., halo,—C₁₋₆alkyl, —CF₃, —C₂₋₆alkynyl, —O—C₁₋₆alkyl, —OCF₃,—C₁₋₄alkylene-O—C₁₋₄alkyl, —C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl,—C₀₋₆alkylene-OH, —CN, —C₀₋₂alkylene-COOH, —CHO, —C(O)—C₁₋₆alkyl,—C(O)O—C₁₋₄alkyl, —CH₂SH, —S—C₁₋₆alkyl, —C₁₋₄alkylene-S—C₁₋₄alkyl,—SO₂—C₁₋₆alkyl, —SO₂NRR, —NHSO₂R, —C₀₋₁alkylene-NRR, —NHC(O)—C₁₋₆alkyl,—C(O)NRR, and —NO₂.

Exemplary halo groups include fluoro, chloro, bromo, and iodo. Exemplary—C₁₋₆alkyl groups include —CH₃, —CH₂CH₃, and —CH(CH₃)₂. Exemplary—C₂₋₆alkynyl groups include —CH═CH₂. Exemplary —O—C₁₋₆alkyl groupsinclude —OCH₃, —O—CH₂CH₃, and —OCH(CH₃)₂. Exemplary—C₁₋₄alkylene-O—C₁₋₄alkyl groups include —CH₂—OCH₃ and —CH₂—OCH₂CH₃.Exemplary —C₀₋₁alkylene-phenyl groups include phenyl and benzyl.Exemplary —O—C₀₋₃alkylene-phenyl groups include —O-phenyl and —O-benzyl.Exemplary —C₀₋₆alkylene-OH groups include —OH and —CH₂OH. Exemplary—C₀₋₂alkylene-COOH groups include —COOH. Exemplary —C(O)—C₁₋₆alkylgroups include —C(O)CH₃ and —C(O)CH₂CH₃. Exemplary —C(O)O—C₁₋₄alkylgroups include —C(O)OCH₃, —C(O)OCH₂CH₃, and —C(O)OCH(CH₃)₂. Exemplary—S—C₁₋₆alkyl groups include —SCH₃. Exemplary —C₁₋₄alkylene-S—C₁₋₄alkylgroups include —CH₂—S—CH₃. Exemplary —SO₂—C₁₋₆alkyl groups include—SO₂CH₃. Exemplary —SO₂NRR groups include —SO₂NH₂ and —SO₂N(CH₃)₂.Exemplary —NHSO₂R groups include —NHSO₂H and —NHSO₂CH₃. Exemplary—C₀₋₁alkylene-NRR groups include —NH₂, —N(CH₃)₂, —CH₂NH(CH₂CH₃), and—CH₂N(CH₃)(CH₂CH₃). Exemplary —NHC(O)—C₁₋₆alkyl groups include—NHC(O)CH₃ and —NHC(O)CH₂CH₃. Exemplary —C(O)NRR groups include —CONH₂,—CONH(CH₂CH₃), and —C(O)N(CH₃)CH₂CH₃.

In one embodiment, R²⁻⁶ are independently hydrogen, halo, —C₁₋₆alkyl,—O—C₁₋₆alkyl, —C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl, or —CN. Inone particular embodiment, R² is hydrogen, halo (e.g., fluoro, chloro,bromo or iodo), —C₁₋₆alkyl (e.g., methyl, ethyl or isopropyl),—O—C₁₋₆alkyl (e.g., methoxy), —C₀₋₁alkylene-phenyl (e.g., phenyl orbenzyl), —O—C₀₋₃alkylene-phenyl (e.g., —O-phenyl or —O-benzyl), or —CN.In one embodiment, R³ is hydrogen, halo (e.g., fluoro, chloro, bromo oriodo), —C₁₋₆alkyl (e.g., methyl or t-butyl), or —O—C₁₋₆alkyl (e.g.,methoxy). In yet another embodiment, R⁴ is hydrogen, halo (e.g., fluoroor chloro), or —C₁₋₆alkyl (e.g., methyl). In another embodiment, R⁵ ishydrogen or halo (e.g., fluoro, chloro, or bromo). In yet anotherembodiment, R⁶ is hydrogen, halo (e.g., chloro), or —C₁₋₆alkyl (e.g.,isopropyl).

In one particular embodiment, the compounds of the invention haveformula II:

where: a, R¹, and R³ through R⁶ are as defined for formula I; or apharmaceutically acceptable salt thereof; with the proviso that thecompound of formula II is not1-[2-(2,3-difluorophenoxymethyl)phenyl]piperazine. In one specificembodiment of formula II: a is 0; and R³⁻⁶ are independently hydrogen,halo, —C₁₋₆alkyl, —O—C₁₋₆alkyl, —C₀₋₁alkylene-phenyl,—O—C₀₋₃alkylene-phenyl, or —CN.

In one particular embodiment, the compounds of the invention haveformula III:

where R²⁻⁶ are independently hydrogen, halo, —C₁₋₆alkyl, —O—C₁₋₆alkyl,—C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl, or —CN; or apharmaceutically acceptable salt thereof; with the proviso that thecompound of formula III is not 1-(2-phenoxymethylphenyl)piperazine,1-[2-(3-fluorophenoxymethyl)phenyl]piperazine, or1-[2-(2,3-difluorophenoxymethyl)phenyl]piperazine.

General Synthetic Procedures

Compounds of the invention can be prepared from readily availablestarting materials using the following general methods, the proceduresset forth in the Examples, or by using other methods, reagents, andstarting materials that are known to those skilled in the art. Althoughthe following procedures may illustrate a particular embodiment of theinvention, it is understood that other embodiments of the invention canbe similarly prepared using the same or similar methods or by usingother methods, reagents and starting materials known to those ofordinary skill in the art. It will also be appreciated that wheretypical or preferred process conditions (i.e., reaction temperatures,times, mole ratios of reactants, solvents, pressures, etc.) are given,other process conditions can also be used unless otherwise stated. Whileoptimum reaction conditions will typically vary depending on variousreaction parameters such as the particular reactants, solvents andquantities used, those of ordinary skill in the art can readilydetermine suitable reaction conditions using routine optimizationprocedures.

Additionally, as will be apparent to those skilled in the art,conventional protecting groups may be necessary or desired to preventcertain functional groups from undergoing undesired reactions. Thechoice of a suitable protecting group for a particular functional groupas well as suitable conditions and reagents for protection anddeprotection of such functional groups are well-known in the art.Protecting groups other than those illustrated in the proceduresdescribed herein may be used, if desired. For example, numerousprotecting groups, and their introduction and removal, are described inT. W. Greene and G. M. Wuts, Protecting Groups in Organic Synthesis,Fourth Edition, Wiley, New York, 2006, and references cited therein.

More particularly, in the schemes below, P represents an“amino-protecting group,” a term used herein to mean a protecting groupsuitable for preventing undesired reactions at an amino group.Representative amino-protecting groups include, but are not limited to,t-butoxycarbonyl (Boc), trityl (Tr), benzyloxycarbonyl (Cbz),9-fluorenylmethoxycarbonyl (Fmoc), formyl, benzyl, and the like.Standard deprotection techniques and reagents such as TFA in DCM or HClin 1,4-dioxane, methanol, or ethanol, are used to remove protectinggroups, when present. For example, a Boc group can be removed using anacidic reagent such as hydrochloric acid, trifluoroacetic acid, and thelike; while a Cbz group can be removed by employing catalytichydrogenation conditions such as H₂ (1 atm), 10% Pd/C in an alcoholicsolvent. The schemes are illustrated with Boc as the protecting group.

Suitable inert diluents or solvents for use in these schemes include, byway of illustration and not limitation, tetrahydrofuran (THF),acetonitrile, N,N-dimethylformamide (DMF), dimethylsulfoxide (DMSO),toluene, dichloromethane (DCM), chloroform (CHCl₃), and the like.

All reactions are typically conducted at a temperature within the rangeof about −78° C. to about 110° C., for example at room temperature.Reactions may be monitored by use of thin layer chromatography (TLC),high performance liquid chromatography (HPLC), and/or LCMS untilcompletion. Reactions may be complete in minutes, may take hours,typically from 1-2 hours and up to 48 hours, or days, such as up to 3-4days. Upon completion, the resulting mixture or reaction product may befurther treated in order to obtain the desired product. For example, theresulting mixture or reaction product may be subjected to one or more ofthe following procedures: dilution (for example with saturated NaHCO₃);extraction (for example, with ethyl acetate, CHCl₃, DCM, aqueous HCl);washing (for example, with DCM, saturated aqueous NaCl, or saturatedaqueous NaHCO₃); drying (for example, over MgSO₄ or Na₂SO₄, or invacuo); filtration; being concentrated (for example, in vacuo); beingredissolved (for example in a 1:1 acetic acid:H₂O solution); and/orpurification (for example by preparative HPLC, reverse phase preparativeHPLC, or crystallization).

By way of illustration, compounds of the invention can be prepared bythe following scheme which is detailed in the examples.

The starting material 1, for example,4-(2-carboxyphenyl)piperidine-1-carboxylic acid t-butyl ester (P=Boc),is commercially available, and undergoes a borane reduction to formcompound 2. Suitable reduction reagents include borane dimethyl sulfidecomplex ((BH₃·DMS), 9-borabicyclo[3.3.1]nonane, borane1,2-bis(t-butylthio)ethane complex, borane t-butylamine complex, boranedi(t-butyl)phosphine complex, borane-tetrahydrofuran complex ((BH₃·THF)and so forth. The next step involves coupling compound 2 with thedesired substituted phenyl under modified Ullmann reaction conditions,where X is —O—SO₂—CF₃ (triflate) or iodide. The modified Ullmannreaction is typically conducted in the presence of a copper(I)iodide/1,10-phenanthroline catalyst and a base such as cesium carbonate,in an appropriate solvent such as toluene or DMF. The resulting productis then deprotected to yield a compound of the invention. This finalstep is conducted under standard deprotection conditions, which willvary depending upon the protecting group used. For example, removal ofthe BOC group can be done using trifluoroacetic acid anddichloromethane, HCl and ethanol, or HCL and dioxane. The triflatecompound is either commercially available, or is readily synthesized bytechniques that are well known in the art.

If desired, pharmaceutically acceptable salts of the compounds offormula I can be prepared by contacting the free acid or base form of acompound of formula I with a pharmaceutically acceptable base or acid.

Certain of the intermediates described herein are believed to be noveland accordingly, such compounds are provided as further aspects of theinvention including, for example, compounds of formula IV:

or a salt thereof, where P represents an amino-protecting group,particularly t-butoxycarbonyl (Boc) where a, and R¹⁻⁶ are as defined forformulas I, II, or III. In one embodiment of the invention, compounds ofthe invention can be prepared by deprotecting compounds of formula IV toprovide compounds of formula I, II, or III, or a pharmaceuticallyacceptable salt thereof.

Further details regarding specific reaction conditions and otherprocedures for preparing representative compounds of the invention orintermediates thereof are described in the Examples set forth below.

Utility

Compounds of the invention possess serotonin and norepinephrine reuptakeinhibitory activity. Thus, these compounds are expected to havetherapeutic utility as combined serotonin and norepinephrine reuptakeinhibitors (SNRIs). In one embodiment, compounds of the inventionpossess equal or approximately equal serotonin reuptake inhibitoryactivity and norepinephrine reuptake inhibitory activity.

The inhibition constant (K_(i)) of a compound is the concentration ofligand in a radioligand binding inhibition assay that would occupy 50%of the transporters if no radioligand were present. K_(i) values can bedetermined from radioligand binding studies with ³H-nisoxetine (for thenorepinephrine transporter, NET) and ³H-citalopram (for the serotonintransporter, SERT), as described in Assay 1. These K_(i) values arederived from IC₅₀ values in the binding assay using the Cheng-Prusoffequation and the K_(d) of the radioligand (Cheng & Prusoff (1973)Biochem. Pharmacol. 22(23):3099-3108). Functional IC₅₀ values can bedetermined in the functional inhibition of uptake assays described inAssay 2. These IC₅₀ values can be converted to K_(i) values using theCheng-Prusoff equation and the K_(m) of the transmitter for thetransporter. It is noted however, that the uptake assay conditionsdescribed in Assay 2 are such that the IC₅₀ values are very close to theK_(i) values, should a mathematical conversion be desired, since theneurotransmitter concentration (5-HT or NE) used in the assay is wellbelow its K_(m) for the respective transporter.

One measure of the affinity of a compound for SERT or NET is theinhibitory constant (pK_(i)) for binding to the transporter. The pK_(i)value is the negative logarithm to base 10 of the K_(i). Compounds ofthe invention of particular interest are those having a pK_(i) at SERTgreater than or equal to about 5.0; and in another embodiment greaterthan or equal to about 7.0. Compounds of the invention of particularinterest also include those having a pK_(i) at NET greater than or equalto about 5.0; and in another embodiment greater than or equal to about7.0. In another embodiment, compounds of interest have a pK_(i) at bothSERT and NET greater than or equal to about 7.0. In another embodiment,compounds of interest have balanced SERT and NET activity, i.e., havethe same pK_(i) value at both SERT and NET±0.5. Such values can bedetermined by techniques that are well known in the art, as well as inthe assays described herein.

In one embodiment, compounds of the invention exhibit a NET pK_(i)≧7and: a SERT K_(i)/NET K_(i) in the range of 0.1 to 100; a SERT K_(i)/NETK_(i) in the range of 0.3 to 100; a SERT K_(i)/NET K_(i) in the range of0.3 to 10; or a SERT K_(i)/NET K_(i) in the range of 0.1 to 30.

Another measure of serotonin and norepinephrine reuptake inhibition isthe pIC₅₀ value. In one embodiment, compounds of interest have aserotonin reuptake inhibition pIC₅₀ value of greater than or equal toabout 7.0 and a norepinephrine reuptake inhibition pIC₅₀ value ofgreater than or equal to about 7.0. In one particular embodiment, thecompounds have a serotonin reuptake inhibition pIC₅₀ value of greaterthan or equal to about 7.0 and a norepinephrine reuptake inhibitionpIC₅₀ value of greater than or equal to about 7.0. In one particularembodiment, the compounds of the invention have balanced pIC₅₀ values,i.e., have the same pIC₅₀ value at both SERT and NET±0.5.

In another embodiment, compounds of the invention are selective forinhibition of SERT and NET over the dopamine transporter (DAT). Forexample in this embodiment, compounds of particular interest are thosethat exhibit a binding affinity for SERT and NET that is at least 5times higher than the binding affinity for DAT, or that is at least 10times higher than for DAT, or at least 20 or 30 times higher than forDAT. In another embodiment, the compounds do not exhibit significant DATinhibition. In still another embodiment, the compounds exhibit less than50% inhibition of DAT activity when measured at a concentration of 794nM. Under the assay conditions used, a compound which exhibits ≦50%inhibition would have an estimated pK_(i) value at DAT of ≦6.1.

In still another embodiment, compounds of the invention possess dopaminereuptake inhibitory activity as well as SERT and NET activity. Forexample in this embodiment, compounds of particular interest are thosethat exhibit a pK_(i) at SERT and NET greater than or equal to about7.0, and a pK_(i) at DAT greater than or equal to about 7.0.

It is noted that in some cases, compounds of the invention may possesseither weak serotonin reuptake inhibitory activity or weaknorepinephrine reuptake inhibitory activity. In these cases, those ofordinary skill in the art will recognize that such compounds still haveutility as primarily either a NET inhibitor or a SERT inhibitor,respectively, or will have utility as research tools.

Exemplary assays to determine the serotonin and/or norepinephrinereuptake inhibiting activity of compounds of the invention include byway of illustration and not limitation, assays that measure SERT and NETbinding, for example, as described in Assay 1 and in Tsuruda et al.(2010) Journal of Pharmacological and Toxicological Methods61(2):192-204. In addition, it is useful to understand the level of DATbinding and uptake in an assay such as that described in Assay 1. Usefulsecondary assays include neurotransmitter uptake assays to measureinhibition of serotonin and norepinephrine uptake into cells expressingthe respective human or rat recombinant transporter (hSERT, hNET, orhDAT) as described in Assay 2, and ex vivo radioligand binding andneurotransmitter uptake assays that are used to determine the in vivooccupancy of SERT, NET and DAT in tissue as described in Assay 3. Otherassays that are useful to evaluate pharmacological properties of testcompounds include those listed in Assay 4. Exemplary in vivo assaysinclude the formalin paw test described in Assay 5, which is a reliablepredictor of clinical efficacy for the treatment of neuropathic pain,and the spinal nerve ligation model described in Assay 6. Theaforementioned assays are useful in determining the therapeutic utility,for example, the neuropathic pain relieving activity, of compounds ofthe invention. Other properties and utilities of compounds of theinvention can be demonstrated using various in vitro and in vivo assayswell-known to those skilled in the art.

Compounds of the invention are expected to be useful for the treatmentand/or prevention of medical conditions in which the regulation ofmonoamine transporter function is implicated, in particular thoseconditions mediated by or responsive to the inhibition of serotonin andnorepinephrine reuptake. Thus it is expected that patients sufferingfrom a disease or disorder that is treated by the inhibition of theserotonin and/or the norepinephrine transporter can be treated byadministering a therapeutically effective amount of a serotonin andnorepinephrine reuptake inhibitor of the invention. Such medicalconditions include, by way of example: pain disorders such asneuropathic pain, fibromyalgia, and chronic pain; depressive disorderssuch as major depression; affective disorders such as an anxietydisorder; attention deficit hyperactivity disorder; cognitive disorderssuch as dementia; stress urinary incontinence; chronic low back pain;and osteoarthritis.

The amount of active agent administered per dose or the total amountadministered per day may be predetermined or it may be determined on anindividual patient basis by taking into consideration numerous factors,including the nature and severity of the patient's condition, thecondition being treated, the age, weight, and general health of thepatient, the tolerance of the patient to the active agent, the route ofadministration, pharmacological considerations such as the activity,efficacy, pharmacokinetics and toxicology profiles of the active agentand any secondary agents being administered, and the like. Treatment ofa patient suffering from a disease or medical condition (such asneuropathic pain) can begin with a predetermined dosage or a dosagedetermined by the treating physician, and will continue for a period oftime necessary to prevent, ameliorate, suppress, or alleviate thesymptoms of the disease or medical condition. Patients undergoing suchtreatment will typically be monitored on a routine basis to determinethe effectiveness of therapy. For example, in treating neuropathic pain,a measure of the effectiveness of treatment may involve assessment ofthe patient's quality of life, e.g., improvements in the patient'ssleeping patterns, work attendance, ability to exercise and beambulatory, etc. Pain scales, operating on a point basis, may also beused to help evaluate a patient's pain level. Indicators for the otherdiseases and conditions described herein, are well-known to thoseskilled in the art, and are readily available to the treating physician.Continuous monitoring by the physician will insure that the optimalamount of active agent will be administered at any given time, as wellas facilitating the determination of the duration of treatment. This isof particular value when secondary agents are also being administered,as their selection, dosage, and duration of therapy may also requireadjustment. In this way, the treatment regimen and dosing schedule canbe adjusted over the course of therapy so that the lowest amount ofactive agent that exhibits the desired effectiveness is administeredand, further, that administration is continued only so long as isnecessary to successfully treat the disease or medical condition.

Pain Disorders

SNRIs have been shown to have a beneficial effect on pain such aspainful diabetic neuropathy (duloxetine, Goldstein et al. (2005) Pain116:109-118; venlafaxine, Rowbotham et al. (2004) Pain 110:697-706),fibromyalgia (duloxetine, Russell et al. (2008) Pain 136(3):432-444;milnacipran, Vitton et al. (2004) Human Psychopharmacology 19:S27-S35),and migraine (venlafaxine, Ozyalcin et al. (2005) Headache45(2):144-152). Thus, one embodiment of the invention relates to amethod for treating a pain disorder, comprising administering to apatient a therapeutically effective amount of a compound of theinvention. Typically, the therapeutically effective amount will be theamount that is sufficient to relieve the pain. Exemplary pain disordersinclude, by way of illustration, acute pain, persistent pain, chronicpain, inflammatory pain, and neuropathic pain. More specifically, theseinclude pain associated with or caused by: arthritis; back painincluding chronic low back pain; cancer, including tumor related pain(e.g., bone pain, headache, facial pain or visceral pain) and painassociated with cancer therapy (e.g., post-chemotherapy syndrome,chronic post-surgical pain syndrome and post-radiation syndrome); carpaltunnel syndrome; fibromyalgia; headaches including chronic tensionheadaches; inflammation associated with polymyalgia, rheumatoidarthritis and osteoarthritis; migraine; neuropathic pain includingcomplex regional pain syndrome; overall pain; post-operative pain;shoulder pain; central pain syndromes, including post-stroke pain, andpain associated with spinal cord injuries and multiple sclerosis;phantom limb pain; pain associated with Parkinson's disease; andvisceral pain (e.g., irritable bowel syndrome). Of particular interestis the treatment of neuropathic pain, which includes diabetic peripheralneuropathy (DPN), HIV-related neuropathy, post-herpetic neuralgia (PHN),and chemotherapy-induced peripheral neuropathy. When used to treat paindisorders such as neuropathic pain, compounds of the invention may beadministered in combination with other therapeutic agents, includinganticonvulsants, antidepressants, muscle relaxants, NSAIDs, opioidagonists, opioid antagonists, selective serotonin reuptake inhibitors,sodium channel blockers, and sympatholytics. Exemplary compounds withinthese classes are described herein.

Depressive Disorders

Another embodiment of the invention relates to a method of treating adepressive disorder, comprising administering to a patient atherapeutically effective amount of a compound of the invention.Typically, the therapeutically effective amount will be the amount thatis sufficient to alleviate depression and provide a sense of generalwell-being. Exemplary depressive disorders include, by way ofillustration and not limitation: depression associated with Alzheimer'sdisease, bipolar disorder, cancer, child abuse, infertility, Parkinson'sdisease, postmyocardial infarction, and psychosis; dysthymia; grumpy orirritable old man syndrome; induced depression; major depression;pediatric depression; postmenopausal depression; post partum depression;recurrent depression; single episode depression; and subsyndromalsymptomatic depression. Of particular interest is the treatment of majordepression. When used to treat depressive disorders, compounds of theinvention may be administered in combination with other therapeuticagents, including antidepressants and dual serotonin-norepinephrinereuptake inhibitors. Exemplary compounds within these classes aredescribed herein.

Affective Disorders

Another embodiment of the invention relates to a method of treating anaffective disorder, comprising administering to a patient atherapeutically effective amount of a compound of the invention.Exemplary affective disorders include, by way of illustration and notlimitation: anxiety disorders such as general anxiety disorder; avoidantpersonality disorder; eating disorders such as anorexia nervosa, bulimianervosa and obesity; obsessive compulsive disorder; panic disorder;personality disorders such as avoidant personality disorder andattention deficit hyperactivity disorder (ADHD); post-traumatic stresssyndrome; phobias such as agoraphobia, as well as simple and otherspecific phobias, and social phobia; premenstrual syndrome; psychoticdisorders, such as schizophrenia and mania; seasonal affective disorder;sexual dysfunction, including premature ejaculation, male impotence, andfemale sexual dysfunction such as female sexual arousal disorder; socialanxiety disorder; and substance abuse disorders, including chemicaldependencies such as addictions to alcohol, benzodiazepines, cocaine,heroin, nicotine and phenobarbital, as well as withdrawal syndromes thatmay arise from these dependencies. When used to treat affectivedisorders, compounds of the invention may be administered in combinationwith other therapeutic agents, including antidepressants. Exemplarycompounds within these classes are described herein.

Atomoxetine, which is 10-fold NET selective, is approved for attentiondeficit hyperactivity disorder (ADHD) therapy, and clinical studies haveshown that the SNRT, venlafaxine, can also have a beneficial effect intreating ADHD (Mukaddes et al. (2002) Eur. Neuropsychopharm. 12(Supp3):421). Thus, the compounds of the invention are also expected to beuseful in methods for treating attention deficit hyperactivity disorderby administering to a patient a therapeutically effective amount of acompound of the invention. When used to treat depression, compounds ofthe invention may be administered in combination with other therapeuticagents, including antidepressants. Exemplary compounds within theseclasses are described herein.

Cognitive Disorders

Another embodiment of the invention relates to a method of treating acognitive disorder, comprising administering to a patient atherapeutically effective amount of a compound of the invention.Exemplary cognitive disorders include, by way of illustration and notlimitation: dementia, which includes degenerative dementia (e.g.,Alzheimer's disease, Creutzfeldt-Jakob disease, Huntington's chorea,Parkinson's disease, Pick's disease, and senile dementia), vasculardementia (e.g., multi-infarct dementia), and dementia associated withintracranial space occupying lesions, trauma, infections and relatedconditions (including HIV infection), metabolism, toxins, anoxia andvitamin deficiency; and mild cognitive impairment associated withageing, such as age associated memory impairment, amnesiac disorder andage-related cognitive decline. When used to treat cognitive disorders,compounds of the invention may be administered in combination with othertherapeutic agents, including anti-Alzheimer's agents andanti-Parkinson's agents. Exemplary compounds within these classes aredescribed herein.

Other Disorders

SNRIs have also been shown to be effective for the treatment of stressurinary incontinence (Dmochowski (2003) Journal of Urology 170(4):1259-1263). Thus, another embodiment of the invention relates to amethod for treating stress urinary incontinence, comprisingadministering to a patient a therapeutically effective amount of acompound of the invention. When used to treat stress urinaryincontinence, compounds of the invention may be administered incombination with other therapeutic agents, including anticonvulsants.Exemplary compounds within these classes are described herein.

Duloxetine, an SNRI, is undergoing clinical trials for evaluating itsefficacy in treating chronic fatigue syndrome, and has recently beenshown to be effective in treating fibromyalgia (Russell et al. (2008)Pain 136(3):432-444). The compounds of the invention, due to theirability to inhibit SERT and NET, are also expected to have this utility,and another embodiment of the invention relates to a method for treatingchronic fatigue syndrome, comprising administering to a patient atherapeutically effective amount of a compound of the invention.

Sibutramine, a norepinephrine and dopamine reuptake inhibitor, has beenshown to be useful in treating obesity (Wirth et al. (2001) JAMA286(11):1331-1339). The compounds of the invention, due to their abilityto inhibit NET, are also expected to have this utility, and anotherembodiment of the invention relates to a method for treating obesity,comprising administering to a patient a therapeutically effective amountof a compound of the invention.

Desvenlafaxine, an SNRI, has been shown to relieve vasomotor symptomsassociated with menopause (Deecher et al. (2007) Endocrinology148(3):1376-1383). The compounds of the invention, due to their abilityto inhibit SERT and NET, are also expected to have this utility, andanother embodiment of the invention relates to a method for treatingvasomotor symptoms associated with menopause, comprising administeringto a patient a therapeutically effective amount of a compound of theinvention.

Research Tools

Since compounds of the invention possess both serotonin reuptakeinhibition activity and norepinephrine reuptake inhibition activity,such compounds are also useful as research tools for investigating orstudying biological systems or samples having serotonin ornorepinephrine transporters. Any suitable biological system or samplehaving serotonin and/or norepinephrine transporters may be employed insuch studies which may be conducted either in vitro or in vivo.Representative biological systems or samples suitable for such studiesinclude, but are not limited to, cells, cellular extracts, plasmamembranes, tissue samples, isolated organs, mammals (such as mice, rats,guinea pigs, rabbits, dogs, pigs, humans, and so forth), and the like,with mammals being of particular interest. In one particular embodimentof the invention, serotonin reuptake in a mammal is inhibited byadministering a serotonin reuptake-inhibiting amount of a compound ofthe invention. In another particular embodiment, norepinephrine reuptakein a mammal is inhibited by administering a norepinephrinereuptake-inhibiting amount of a compound of the invention. Compounds ofthe invention can also be used as research tools by conductingbiological assays using such compounds.

When used as a research tool, a biological system or sample comprising aserotonin transporter and/or a norepinephrine transporter is typicallycontacted with a serotonin reuptake-inhibiting or norepinephrinereuptake-inhibiting amount of a compound of the invention. After thebiological system or sample is exposed to the compound, the effects ofinhibiting serotonin reuptake and/or norepinephrine reuptake aredetermined using conventional procedures and equipment. Exposureencompasses contacting cells or tissue with the compound, administeringthe compound to a mammal, for example by i.p. or i.v. administration,and so forth. This determining step may comprise measuring a response,i.e., a quantitative analysis or may comprise an observation, i.e., aqualitative analysis. Measuring a response involves, for example,determining the effects of the compound on the biological system orsample using conventional procedures and equipment, such as serotoninand norepinephrine reuptake assays. The assay results can be used todetermine the activity level as well as the amount of compound necessaryto achieve the desired result, i.e., a serotonin reuptake-inhibiting anda norepinephrine reuptake-inhibiting amount.

Additionally, compounds of the invention can be used as research toolsfor evaluating other chemical compounds, and thus are also useful inscreening assays to discover, for example, new compounds having bothserotonin reuptake-inhibiting activity and norepinephrinereuptake-inhibiting activity. In this manner, compounds of the inventionare used as standards in an assay to allow comparison of the resultsobtained with a test compound and with compounds of the invention toidentify those test compounds that have about equal or superiorreuptake-inhibiting activity, if any. For example, reuptake data for atest compound or a group of test compounds is compared to the reuptakedata for a compound of the invention to identify those test compoundsthat have the desired properties, e.g., test compounds havingreuptake-inhibiting activity about equal or superior to a compound ofthe invention, if any. This aspect of the invention includes, asseparate embodiments, both the generation of comparison data (using theappropriate assays) and the analysis of the test data to identify testcompounds of interest. Thus, a test compound can be evaluated in abiological assay, by a method comprising the steps of: (a) conducting abiological assay with a test compound to provide a first assay value;(b) conducting the biological assay with a compound of the invention toprovide a second assay value; wherein step (a) is conducted eitherbefore, after or concurrently with step (b); and (c) comparing the firstassay value from step (a) with the second assay value from step (b).Exemplary biological assays include serotonin and norepinephrinereuptake assays.

Pharmaceutical Compositions and Formulations

Compounds of the invention are typically administered to a patient inthe form of a pharmaceutical composition or formulation. Suchpharmaceutical compositions may be administered to the patient by anyacceptable route of administration including, but not limited to, oral,rectal, vaginal, nasal, inhaled, topical (including transdermal) andparenteral modes of administration. Further, the compounds of theinvention may be administered, for example orally, in multiple doses perday (e.g., twice, three times or four times daily), in a single dailydose, in a twice daily dose, in a single weekly dose, and so forth. Itwill be understood that any form of the compounds of the invention,(i.e., free base, pharmaceutically acceptable salt, solvate, etc.) thatis suitable for the particular mode of administration can be used in thepharmaceutical compositions discussed herein.

Accordingly, in one embodiment, the invention relates to apharmaceutical composition comprising a pharmaceutically acceptablecarrier and a compound of the invention. The compositions may containother therapeutic and/or formulating agents if desired. When discussingcompositions, the “compound of the invention” may also be referred toherein as the “active agent,” to distinguish it from other components ofthe formulation, such as the carrier. Thus, it is understood that theterm “active agent” includes compounds of formula I as well aspharmaceutically acceptable salts and solvates of that compound.

Pharmaceutical compositions of the invention typically contain atherapeutically effective amount of a compound of the invention. Thoseskilled in the art will recognize, however, that a pharmaceuticalcomposition may contain more than a therapeutically effective amount,i.e., bulk compositions, or less than a therapeutically effectiveamount, i.e., individual unit doses designed for multiple administrationto achieve a therapeutically effective amount. Typically, thecomposition will contain from about 0.01-95 wt % of active agent,including, from about 0.01-30 wt %, such as from about 0.01-10 wt %,with the actual amount depending upon the formulation itself, the routeof administration, the frequency of dosing, and so forth. In oneembodiment, a composition suitable for an oral dosage form, for example,may contain about 5-70 wt %, or from about 10-60 wt % of active agent.In one exemplary embodiment, a pharmaceutical composition contains fromabout 1 to 20 mg of active agent, including from about 1 to 15 mg ofactive agent and from about 1 to 10 mg of active agent. In anotherexemplary embodiment, a pharmaceutical composition contains from about 5to 20 mg of active agent, including from about 7.5 to 15 mg of activeagent. For example the active agent may be formulated in 1 mg and 10 mgunit doses.

Any conventional carrier or excipient may be used in the pharmaceuticalcompositions of the invention. The choice of a particular carrier orexcipient, or combinations of carriers or excipients, will depend on themode of administration being used to treat a particular patient or typeof medical condition or disease state. In this regard, the preparationof a suitable composition for a particular mode of administration iswell within the scope of those skilled in the pharmaceutical arts.Additionally, carriers or excipients used in such compositions arecommercially available. By way of further illustration, conventionalformulation techniques are described in Remington: The Science andPractice of Pharmacy, 20^(th) Edition, Lippincott Williams & White,Baltimore, Md. (2000); and H. C. Ansel et al., Pharmaceutical DosageForms and Drug Delivery Systems, 7^(th) Edition, Lippincott Williams &White, Baltimore, Md. (1999).

Representative examples of materials which can serve as pharmaceuticallyacceptable carriers include, but are not limited to, the following:sugars, such as lactose, glucose and sucrose; starches, such as cornstarch and potato starch; cellulose, such as microcrystalline cellulose,and its derivatives, such as sodium carboxymethyl cellulose, ethylcellulose and cellulose acetate; powdered tragacanth; malt; gelatin;talc; excipients, such as cocoa butter and suppository waxes; oils, suchas peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil,corn oil and soybean oil; glycols, such as propylene glycol; polyols,such as glycerin, sorbitol, mannitol and polyethylene glycol; esters,such as ethyl oleate and ethyl laurate; agar; buffering agents, such asmagnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-freewater; isotonic saline; Ringer's solution; ethyl alcohol; phosphatebuffer solutions; compressed propellant gases, such aschlorofluorocarbons and hydrofluorocarbons; and other non-toxiccompatible substances employed in pharmaceutical compositions.

Pharmaceutical compositions are typically prepared by thoroughly andintimately mixing or blending the active agent with a pharmaceuticallyacceptable carrier and one or more optional ingredients. The resultinguniformly blended mixture may then be shaped or loaded into tablets,capsules, pills, canisters, cartridges, dispensers, and the like, usingconventional procedures and equipment.

In one embodiment, the pharmaceutical compositions are suitable for oraladministration. One exemplary dosing regimen would be an oral dosageform administered once or twice daily. Suitable compositions for oraladministration may be in the form of capsules, tablets, pills, lozenges,cachets, dragees, powders, granules; solutions or suspensions in anaqueous or non-aqueous liquid; oil-in-water or water-in-oil liquidemulsions; elixirs or syrups; and the like; each containing apredetermined amount of the active agent.

When intended for oral administration in a solid dosage form (i.e., ascapsules, tablets, pills, and the like), the composition will typicallycomprise the active agent and one or more pharmaceutically acceptablecarriers, such as sodium citrate or dicalcium phosphate. Solid dosageforms may also comprise: fillers or extenders, such as starches,microcrystalline cellulose, lactose, sucrose, glucose, mannitol, and/orsilicic acid; binders, such as carboxymethylcellulose, alginates,gelatin, polyvinyl pyrrolidone, sucrose and/or acacia; humectants, suchas glycerol; disintegrating agents, such as agar-agar, calciumcarbonate, potato or tapioca starch, alginic acid, certain silicates,and/or sodium carbonate; solution retarding agents, such as paraffin;absorption accelerators, such as quaternary ammonium compounds; wettingagents, such as cetyl alcohol and/or glycerol monostearate; absorbents,such as kaolin and/or bentonite clay; lubricants, such as talc, calciumstearate, magnesium stearate, solid polyethylene glycols, sodium laurylsulfate, and/or mixtures thereof; coloring agents; and buffering agents.

Release agents, wetting agents, coating agents, sweetening, flavoringand perfuming agents, preservatives and antioxidants may also be presentin the pharmaceutical compositions. Exemplary coating agents fortablets, capsules, pills and like, include those used for entericcoatings, such as cellulose acetate phthalate, polyvinyl acetatephthalate, hydroxypropyl methylcellulose phthalate, methacrylicacid-methacrylic acid ester copolymers, cellulose acetate trimellitate,carboxymethyl ethyl cellulose, hydroxypropyl methyl cellulose acetatesuccinate, and the like. Examples of pharmaceutically acceptableantioxidants include: water-soluble antioxidants, such as ascorbic acid,cysteine hydrochloride, sodium bisulfate, sodium metabisulfate, sodiumsulfite, and the like; oil-soluble antioxidants, such as ascorbylpalmitate, butylated hydroxyanisole, butylated hydroxytoluene, lecithin,propyl gallate, alpha-tocopherol, and the like; and metal-chelatingagents, such as citric acid, ethylenediamine tetraacetic acid, sorbitol,tartaric acid, phosphoric acid, and the like.

Compositions may also be formulated to provide slow or controlledrelease of the active agent using, by way of example, hydroxypropylmethyl cellulose in varying proportions or other polymer matrices,liposomes and/or microspheres. In addition, the pharmaceuticalcompositions of the invention may contain opacifying agents and may beformulated so that they release the active agent only, orpreferentially, in a certain portion of the gastrointestinal tract,optionally, in a delayed manner. Examples of embedding compositionswhich can be used include polymeric substances and waxes. The activeagent can also be in micro-encapsulated form, if appropriate, with oneor more of the above-described excipients.

Suitable liquid dosage forms for oral administration include, by way ofillustration, pharmaceutically acceptable emulsions, microemulsions,solutions, suspensions, syrups and elixirs. Liquid dosage formstypically comprise the active agent and an inert diluent, such as, forexample, water or other solvents, solubilizing agents and emulsifiers,such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethylacetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyleneglycol, oils (e.g., cottonseed, groundnut, corn, germ, olive, castor andsesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycolsand fatty acid esters of sorbitan, and mixtures thereof. Suspensions maycontain suspending agents such as, for example, ethoxylated isostearylalcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystallinecellulose, aluminium metahydroxide, bentonite, agar-agar and tragacanth,and mixtures thereof.

When intended for oral administration, the pharmaceutical compositionsof the invention may be packaged in a unit dosage form. The term “unitdosage form” refers to a physically discrete unit suitable for dosing apatient, i.e., each unit containing a predetermined quantity of theactive agent calculated to produce the desired therapeutic effect eitheralone or in combination with one or more additional units. For example,such unit dosage forms may be capsules, tablets, pills, and the like.

In another embodiment, the compositions of the invention are suitablefor inhaled administration, and will typically be in the form of anaerosol or a powder. Such compositions are generally administered usingwell-known delivery devices, such as a nebulizer, dry powder, ormetered-dose inhaler. Nebulizer devices produce a stream of highvelocity air that causes the composition to spray as a mist that iscarried into a patient's respiratory tract. An exemplary nebulizerformulation comprises the active agent dissolved in a carrier to form asolution, or micronized and combined with a carrier to form a suspensionof micronized particles of respirable size. Dry powder inhalersadminister the active agent as a free-flowing powder that is dispersedin a patient's air-stream during inspiration. An exemplary dry powderformulation comprises the active agent dry-blended with an excipientsuch as lactose, starch, mannitol, dextrose, polylactic acid,polylactide-co-glycolide, and combinations thereof. Metered-doseinhalers discharge a measured amount of the active agent usingcompressed propellant gas. An exemplary metered-dose formulationcomprises a solution or suspension of the active agent in a liquefiedpropellant, such as a chlorofluorocarbon or hydrofluoroalkane. Optionalcomponents of such formulations include co-solvents, such as ethanol orpentane, and surfactants, such as sorbitan trioleate, oleic acid,lecithin, and glycerin. Such compositions are typically prepared byadding chilled or pressurized hydrofluoroalkane to a suitable containercontaining the active agent, ethanol (if present) and the surfactant (ifpresent). To prepare a suspension, the active agent is micronized andthen combined with the propellant. Alternatively, a suspensionformulation can be prepared by spray drying a coating of surfactant onmicronized particles of the active agent. The formulation is then loadedinto an aerosol canister, which forms a portion of the inhaler.

Compounds of the invention can also be administered parenterally (e.g.,by subcutaneous, intravenous, intramuscular, or intraperitonealinjection). For such administration, the active agent is provided in asterile solution, suspension, or emulsion. Exemplary solvents forpreparing such formulations include water, saline, low molecular weightalcohols such as propylene glycol, polyethylene glycol, oils, gelatin,fatty acid esters such as ethyl oleate, and the like. A typicalparenteral formulation is a sterile pH 4-7 aqueous solution of theactive agent. Parenteral formulations may also contain one or moresolubilizers, stabilizers, preservatives, wetting agents, emulsifiers,and dispersing agents. These formulations may be rendered sterile by useof a sterile injectable medium, a sterilizing agent, filtration,irradiation, or heat.

Compounds of the invention can also be administered transdermally usingknown transdermal delivery systems and excipients. For example, thecompound can be admixed with permeation enhancers, such as propyleneglycol, polyethylene glycol monolaurate, azacycloalkan-2-ones, and thelike, and incorporated into a patch or similar delivery system.Additional excipients including gelling agents, emulsifiers and buffers,may be used in such transdermal compositions if desired.

If desired, compounds of the invention may be administered incombination with one or more other therapeutic agents. Thus, in oneembodiment, compositions of the invention may optionally contain otherdrugs that are co-administered with a compound of the invention. Forexample, the composition may further comprise one or more drugs (alsoreferred to as “secondary agents(s)”) selected from the group ofanti-Alzheimer's agents, anticonvulsants (antiepileptics),antidepressants, anti-Parkinson's agents, dual serotonin-norepinephrinereuptake inhibitors (SNRIs), non-steroidal anti-inflammatory agents(NSAIDs), norepinephrine reuptake inhibitors, opioid agonists (opioidanalgesics), opioid antagonists, selective serotonin reuptakeinhibitors, sodium channel blockers, sympatholytics, and combinationsthereof. Numerous examples of such therapeutic agents are well known inthe art, and examples are described herein. By combining a compound ofthe invention with a secondary agent, triple therapy can be achieved,i.e., serotonin reuptake inhibitory activity, norepinephrine reuptakeinhibitory activity, and activity associated with the secondary agent(e.g., antidepressant activity), using only two active components. Sincepharmaceutical compositions containing two active components aretypically easier to formulate than compositions containing three activecomponents, such two-component compositions provide a significantadvantage over compositions containing three active components.Accordingly, in yet another aspect of the invention, a pharmaceuticalcomposition comprises a compound of the invention, a second activeagent, and a pharmaceutically acceptable carrier. Third, fourth, etc.,active agents may also be included in the composition. In combinationtherapy, the amount of compound of the invention that is administered,as well as the amount of secondary agents, may be less than the amounttypically administered in monotherapy.

A compound of the invention may be either physically mixed with thesecond active agent to form a composition containing both agents; oreach agent may be present in separate and distinct compositions whichare administered to the patient simultaneously or sequentially. Forexample, a compound of the invention can be combined with a secondactive agent using conventional procedures and equipment to form acombination of active agents comprising a compound of the invention anda second active agent. Additionally, the active agents may be combinedwith a pharmaceutically acceptable carrier to form a pharmaceuticalcomposition comprising a compound of the invention, a second activeagent and a pharmaceutically acceptable carrier. In this embodiment, thecomponents of the composition are typically mixed or blended to create aphysical mixture. The physical mixture is then administered in atherapeutically effective amount using any of the routes describedherein.

Alternatively, the active agents may remain separate and distinct beforeadministration to the patient. In this embodiment, the agents are notphysically mixed together before administration but are administeredsimultaneously or at separate times as separate compositions. Suchcompositions can be packaged separately or may be packaged together in akit. When administered at separate times, the secondary agent willtypically be administered less than 24 hours after administration of thecompound of the invention, ranging anywhere from concurrent withadministration of the compound of the invention to about 24 hourspost-dose. This is also referred to as sequential administration. Thus,a compound of the invention can be orally administered simultaneously orsequentially with another active agent using two tablets, with onetablet for each active agent, where sequential may mean beingadministered immediately after administration of the compound of theinvention or at some predetermined time later (e.g., one hour later orthree hours later). Alternatively, the combination may be administeredby different routes of administration, i.e., one orally and the other byinhalation.

In one embodiment, the kit comprises a first dosage form comprising acompound of the invention and at least one additional dosage formcomprising one or more of the secondary agents set forth herein, inquantities sufficient to carry out the methods of the invention. Thefirst dosage form and the second (or third, etc.) dosage form togethercomprise a therapeutically effective amount of active agents for thetreatment or prevention of a disease or medical condition in a patient.

Secondary agent(s), when included, are present in a therapeuticallyeffective amount, i.e., are typically administered in an amount thatproduces a therapeutically beneficial effect when co-administered with acompound of the invention. The secondary agent can be in the form of apharmaceutically acceptable salt, solvate, optically pure stereoisomer,and so forth. Thus, secondary agents listed below are intended toinclude all such forms, and are commercially available or can beprepared using conventional procedures and reagents.

Representative anti-Alzheimer's agents include, but are not limited to:donepezil, galantamine, memantine, rivastigmine, selegiline, tacrine,and combinations thereof.

Representative anticonvulsants (antiepileptics) include, but are notlimited to: acetazolamide, albutoin, 4-amino-3-hydroxybutyric acid,beclamide, carbamazepine, cinromide, clomethiazole, clonazepam,diazepam, dimethadione, eterobarb, ethadione, ethosuximide, ethotoin,felbamate, fosphenyloin, gabapentin, lacosamide, lamotrigine, lorazepam,magnesium bromide, magnesium sulfate, mephenyloin, mephobarbital,methsuximide, midazolam, nitrazepam, oxazepam, oxcarbazepine,paramethadione, phenacemide, pheneturide, phenobarbital, phensuximide,phenyloin, potassium bromide, pregabalin, primidone, progabide, sodiumbromide, sodium valproate, sulthiame, tiagabine, topiramate,trimethadione, valproic acid, valpromide, vigabatrin, zonisamide, andcombinations thereof. In a particular embodiment, the anticonvulsant isselected from carbamazepine, gabapentin, pregabalin, and combinationsthereof.

Representative antidepressants include, but are not limited to:adinazolam, amitriptyline, clomipramine, desipramine, dothiepin (e.g.,dothiepin hydrochloride), doxepin, imipramine, lofepramine, mirtazapine,nortriptyline, protriptyline, trimipramine, venlafaxine, zimelidine, andcombinations thereof.

Representative anti-Parkinson's agents include, but are not limited to:amantadine, apomorphine, benztropine, bromocriptine, carbidopa,diphenhydramine, entacapone, levodopa, pergolide, pramipexole,ropinirole, selegiline, tolcapone, trihexyphenidyl, and combinationsthereof.

Representative dual serotonin-norepinephrine reuptake inhibitors (SNRIs)include, but are not limited to: bicifadine, desvenlafaxine, duloxetine,milnacipran, nefazodone, venlafaxine, and combinations thereof.

Representative non-steroidal anti-inflammatory agents (NSAIDs) include,but are not limited to: acemetacin, acetaminophen, acetyl salicylicacid, alclofenac, alminoprofen, amfenac, amiprilose, amoxiprin,anirolac, apazone, azapropazone, benorilate, benoxaprofen, bezpiperylon,broperamole, bucloxic acid, carprofen, clidanac, diclofenac, diflunisal,diftalone, enolicam, etodolac, etoricoxib, fenbufen, fenclofenac,fenclozic acid, fenoprofen, fentiazac, feprazone, flufenamic acid,flufenisal, fluprofen, flurbiprofen, furofenac, ibufenac, ibuprofen,indomethacin, indoprofen, isoxepac, isoxicam, ketoprofen, ketorolac,lofemizole, lornoxicam, meclofenamate, meclofenamic acid, mefenamicacid, meloxicam, mesalamine, miroprofen, mofebutazone, nabumetone,naproxen, niflumic acid, nimesulide, nitroflurbiprofen, olsalazine,oxaprozin, oxpinac, oxyphenbutazone, phenylbutazone, piroxicam,pirprofen, pranoprofen, salsalate, sudoxicam, sulfasalazine, sulindac,suprofen, tenoxicam, tiopinac, tiaprofenic acid, tioxaprofen, tolfenamicacid, tolmetin, triflumidate, zidometacin, zomepirac, and combinationsthereof. In a particular embodiment, the NSAID is selected frometodolac, flurbiprofen, ibuprofen, indomethacin, ketoprofen, ketorolac,meloxicam, naproxen, oxaprozin, piroxicam, and combinations thereof. Ina particular embodiment, the NSAID is selected from ibuprofen,indomethacin, nabumetone, naproxen (for example, naproxen sodium), andcombinations thereof.

Representative muscle relaxants include, but are not limited to:carisoprodol, chlorzoxazone, cyclobenzaprine, diflunisal, metaxalone,methocarbamol, and combinations thereof.

Representative norepinephrine reuptake inhibitors include, but are notlimited to: atomoxetine, buproprion and the buproprion metabolitehydroxybuproprion, maprotiline, reboxetine (for example,(S,S)-reboxetine), viloxazine, and combinations thereof. In a particularembodiment, the norepinephrine reuptake inhibitor is selected fromatomoxetine, reboxetine, and combinations thereof.

Representative opioid agonists (opioid analgesics) and antagonistsinclude, but are not limited to: buprenorphine, butorphanol, codeine,dihydrocodeine, fentanyl, hydrocodone, hydromorphone, levallorphan,levorphanol, meperidine, methadone, morphine, nalbuphine, nalmefene,nalorphine, naloxone, naltrexone, nalorphine, oxycodone, oxymorphone,pentazocine, propoxyphene, tramadol, and combinations thereof. Incertain embodiments, the opioid agonist is selected from codeine,dihydrocodeine, hydrocodone, hydromorphone, morphine, oxycodone,oxymorphone, tramadol, and combinations thereof.

Representative selective serotonin reuptake inhibitors (SSRIs) include,but are not limited to: citalopram and the citalopram metabolitedesmethylcitalopram, dapoxetine, escitalopram (e.g., escitalopramoxalate), fluoxetine and the fluoxetine desmethyl metabolitenorfluoxetine, fluvoxamine (e.g., fluvoxamine maleate), paroxetine,sertraline and the sertraline metabolite demethylsertraline, andcombinations thereof. In certain embodiments, the SSRI is selected fromcitalopram, paroxetine, sertraline, and combinations thereof.

Representative sodium channel blockers include, but are not limited to:carbamazepine, fosphenyloin, lamotrignine, lidocaine, mexiletine,oxcarbazepine, phenyloin, and combinations thereof.

Representative sympatholytics include, but are not limited to: atenolol,clonidine, doxazosin, guanethidine, guanfacine, modafinil, phentolamine,prazosin, reserpine, tolazoline (e.g., tolazoline hydrochloride),tamsulosin, and combinations thereof.

The following formulations illustrate representative pharmaceuticalcompositions of the present invention:

Exemplary Hard Gelatin Capsules for Oral Administration

A compound of the invention (50 g), spray-dried lactose (440 g) andmagnesium stearate (10 g) are thoroughly blended. The resultingcomposition is then loaded into hard gelatin capsules (500 mg ofcomposition per capsule).

Alternately, a compound of the invention (20 mg) is thoroughly blendedwith starch (89 mg), microcrystalline cellulose (89 mg) and magnesiumstearate (2 mg). The mixture is then passed through a No. 45 mesh U.S.sieve and loaded into a hard gelatin capsule (200 mg of composition percapsule).

Exemplary Gelatin Capsule Formulation for Oral Administration

A compound of the invention (100 mg) is thoroughly blended withpolyoxyethylene sorbitan monooleate (50 mg) and starch powder (250 mg).The mixture is then loaded into a gelatin capsule (400 mg of compositionper capsule).

Alternately, a compound of the invention (40 mg) is thoroughly blendedwith microcrystalline cellulose (Avicel PH 103; 259.2 mg) and magnesiumstearate (0.8 mg). The mixture is then loaded into a gelatin capsule(Size #1, White, Opaque) (300 mg of composition per capsule).

Exemplary Tablet Formulation for Oral Administration

A compound of the invention (10 mg), starch (45 mg) and microcrystallinecellulose (35 mg) are passed through a No. 20 mesh U.S. sieve and mixedthoroughly. The granules so produced are dried at 50-60° C. and passedthrough a No. 16 mesh U.S. sieve. A solution of polyvinylpyrrolidone (4mg as a 10% solution in sterile water) is mixed with sodiumcarboxymethyl starch (4.5 mg), magnesium stearate (0.5 mg), and talc (1mg), and this mixture is then passed through a No. 16 mesh U.S. sieve.The sodium carboxymethyl starch, magnesium stearate and talc are thenadded to the granules. After mixing, the mixture is compressed on atablet machine to afford a tablet weighing 100 mg.

Alternately, a compound of the invention (250 mg) is thoroughly blendedwith microcrystalline cellulose (400 mg), silicon dioxide fumed (10 mg),and stearic acid (5 mg). The mixture is then compressed to form tablets(665 mg of composition per tablet).

Alternately, a compound of the invention (400 mg) is thoroughly blendedwith cornstarch (50 mg), croscarmellose sodium (25 mg), lactose (120mg), and magnesium stearate (5 mg). The mixture is then compressed toform a single-scored tablet (600 mg of compositions per tablet).

Exemplary Suspension Formulation for Oral Administration

The following ingredients are mixed to form a suspension containing 100mg of active agent per 10 mL of suspension:

Ingredients Amount Compound of the invention 1.0 g Fumaric acid 0.5 gSodium chloride 2.0 g Methyl paraben 0.15 g Propyl paraben 0.05 gGranulated sugar 25.5 g Sorbitol (70% solution) 12.85 g Veegum ® K(magnesium aluminum silicate) 1.0 g Flavoring 0.035 mL Colorings 0.5 mgDistilled water q.s. to 100 mL

Exemplary Injectable Formulation for Administration by Injection

A compound of the invention (0.2 g) is blended with 0.4 M sodium acetatebuffer solution (2.0 mL). The pH of the resulting solution is adjustedto pH 4 using 0.5 N aqueous hydrochloric acid or 0.5 N aqueous sodiumhydroxide, as necessary, and then sufficient water for injection isadded to provide a total volume of 20 mL. The mixture is then filteredthrough a sterile filter (0.22 micron) to provide a sterile solutionsuitable for administration by injection.

Exemplary Compositions for Administration by Inhalation

A compound of the invention (0.2 mg) is micronized and then blended withlactose (25 mg). This blended mixture is then loaded into a gelatininhalation cartridge. The contents of the cartridge are administeredusing a dry powder inhaler, for example.

Alternately, a micronized compound of the invention (10 g) is dispersedin a solution prepared by dissolving lecithin (0.2 g) in demineralizedwater (200 mL). The resulting suspension is spray dried and thenmicronized to form a micronized composition comprising particles havinga mean diameter less than about 1.5 μm. The micronized composition isthen loaded into metered-dose inhaler cartridges containing pressurized1,1,1,2-tetrafluoroethane in an amount sufficient to provide about 10 μgto about 500 μg of the compound of the invention per dose whenadministered by the inhaler.

Alternately, a compound of the invention (25 mg) is dissolved in citratebuffered (pH 5) isotonic saline (125 mL). The mixture is stirred andsonicated until the compound is dissolved. The pH of the solution ischecked and adjusted, if necessary, to pH 5 by slowly adding aqueous 1Nsodium hydroxide. The solution is administered using a nebulizer devicethat provides about 10 μg to about 500 μg of the compound of theinvention per dose.

EXAMPLES

The following Preparations and Examples are provided to illustratespecific embodiments of the invention. These specific embodiments,however, are not intended to limit the scope of the invention in any wayunless specifically indicated.

The following abbreviations have the following meanings unless otherwiseindicated and any other abbreviations used herein and not defined havetheir standard meaning:

-   -   AcOH acetic acid    -   Bn benzyl    -   BSA bovine serum albumin    -   DCM dichloromethane (i.e., methylene chloride)    -   DMEM Dulbecco's Modified Eagle's Medium    -   DMSO dimethylsulfoxide    -   EDTA ethylenediaminetetraacetic acid    -   Et ethyl    -   EtOAc ethyl acetate    -   FBS fetal bovine serum    -   hDAT human dopamine transporter    -   HEPES 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid    -   hNET human norepinephrine transporter    -   hSERT human serotonin transporter    -   5-HT 5-hydroxytryptamine    -   iPr isopropyl    -   Me methyl    -   MeOH methanol    -   NA noradrenaline    -   PBS phosphate buffered saline    -   Ph phenyl    -   tBu t-butyl    -   TFA trifluoroacetic acid    -   THF tetrahydrofuran

Any other abbreviations used herein but not defined have their standard,generally accepted meaning. Unless noted otherwise, all materials, suchas reagents, starting materials and solvents, were purchased fromcommercial suppliers (such as Sigma-Aldrich, Fluka Riedel-de Haen, andthe like) and were used without further purification.

Preparation 1 4-(2-Hydroxymethylphenyl)piperazine-1-carboxylic Acidt-butyl Ester

4-(2-Carboxyphenyl)piperazine-1-carboxylic acid t-butyl ester (3.5 g,11.3 mmol, 1.0 eq.) and THF (91.3 mL) were combined at room temperatureunder nitrogen. Borane-dimethyl sulfide complex (2.00 mL, 22.5 mmol, 2.0eq.) was added dropwise and the mixture was stirred for 5 minutes, thenheated at the reflux temperature for 1.5 hours. The mixture was cooledto room temperature, and the reaction was quenched slowly with MeOH (20ml), then concentrated by rotary evaporation. The material wasazeotroped with MeOH (2×20 ml). The crude product was dissolved in EtOAc(50 ml), and washed with cold 1M HCl (2×25 ml), then NaHCO₃ (2×25 ml).The organic layer was dried over anhydrous Na₂SO₄, filtered, andconcentrated in vacuo to yield the title compound (1.3 g) as a clear oilthat solidified upon sitting.

Example 1 1-[2-(3-Iodophenoxymethyl)phenyl]piperazine

4-(2-Hydroxymethylphenyl)piperazine-1-carboxylic acid t-butyl ester(38.3 mg, 131 μmol, 1.0 eq.), copper(I) iodide (10 mg, 60 μmol, 0.3eq.), trifluoromethanesulfonic acid 3-iodophenyl ester (92 mg, 260 mmol,2.0 eq.), 1,10-phenanthroline (24 mg, 130 μmol, 0.6 eq.) and cesiumcarbonate (170 mg, 520 μmol, 4.4 eq.) in toluene (650 μL) under air. Themixture was heated at 110° C. for 3 days under vigorous stirring.

After the reaction had gone to complete conversion (3 days), the mixturewas diluted with DCM (2 ml) to crash out the salts then filtered. Thefiltrate was then concentrated and redissolved in 2M TFA in DCM (1.0 mL,2.6 mmol) and shaken at room temperature overnight. The crude materialwas then concentrated then the residue was dissolved in 1.5 ml 1:1AcOH/H₂O and an additional 0.3 ml AcOH and filtered. Purification wasperformed by preparative HPLC to yield the title compound as a mono-TFAsalt (4.3 mg, 90% purity). MS m/z: [M+H]⁺ calcd for C₁₇H₁₉₁N₂O, 395.05.found 395.0.

Example 2

Following the procedures described in the examples above, andsubstituting the appropriate starting materials and reagents, compounds2-1 to 2-40, having formula Ia, were also prepared:

(Ia)

MS m/z: [M + H]⁺ Ex. R² R³ R⁴ R⁵ R⁶ Formula calcd found 2-1 F H H H HC₁₇H₁₉FN₂O 287.15 287.2 2-2 Cl H H H H C₁₇H₁₉ClN₂O 303.12 303.0 2-3 I HH H H C₁₇H₁₉IN₂O 395.05 395.0 2-4 Me H H H H C₁₈H₂₂N₂O 283.17 283.2 2-5iPr H H H H C₂₀H₂₆N₂O 311.20 311.2 2-6 Et H H H H C₁₉H₂₄N₂O 297.19 297.22-7 OMe H H H H C₁₈H₂₂N₂O₂ 299.17 299.2 2-8 Ph H H H H C₂₃H₂₄N₂O 345.19345.2 2-9 Bn H H H H C₂₄H₂₆N₂O 359.20 359.2 2-10 OPh H H H H C₂₃H₂₄N₂O₂361.18 361.2 2-11 OBn H H H H C₂₄H₂₆N₂O₂ 375.20 375.2 2-12 H Cl H H HC₁₇H₁₉ClN₂O 303.12 303.2 2-13 H Br H H H C₁₇H₁₉BrN₂O 347.07 347.0 2-14 HMe H H H C₁₈H₂₂N₂O 283.17 283.2 2-15 H tBu H H H C₂₁H₂₈N₂O 325.22 325.22-16 H OMe H H H C₁₈H₂₂N₂O₂ 299.17 299.2 2-17 H H Me H H C₁₈H₂₂N₂O283.17 283.2 2-18 F Cl H H H C₁₇H₁₈ClFN₂O 321.11 321.0 2-19 Cl Cl H H HC₁₇H₁₈Cl₂N₂O 337.08 337.0 2-20 Me F H H H C₁₈H₂₁FN₂O 301.16 301.2 2-21Me Cl H H H C₁₈H₂₁ClN₂O 317.13 317.2 2-22 CN F H H H C₁₈H₁₈FN₃O 312.14312.2 2-23 F H H F H C₁₇H₁₈F₂N₂O 305.14 305.0 2-24 F H H Cl HC₁₇H₁₈ClFN₂O 321.11 321.0 2-25 F H H Br H C₁₇H₁₈BrFN₂O 365.06 365.0 2-26Cl H H Cl H C₁₇H₁₈Cl₂N₂O 337.08 337.0 2-27 Cl H H F H C₁₇H₁₈ClFN₂O321.11 321.0 2-28 Br H H F H C₁₇H₁₈BrFN₂O 365.06 365.0 2-29 Me H H F HC₁₈H₂₁FN₂O 301.16 301.2 2-30 Me H H Cl H C₁₈H₂₁ClN₂O 317.13 317.2 2-31OMe H H Cl H C₁₈H₂₁ClN₂O₂ 333.13 333.2 2-32 CN H H Cl H C₁₈H₁₈ClN₃O328.11 328.0 2-33 H F F H H C₁₇H₁₈F₂N₂O 305.14 305.2 2-34 H F Cl H HC₁₇H₁₈ClFN₂O 321.11 321.0 2-35 H Cl Cl H H C₁₇H₁₈Cl₂N₂O 337.08 337.02-36 H F H F H C₁₇H₁₈F₂N₂O 305.14 305.2 2-37 H Cl H Cl H C₁₇H₁₈Cl₂N₂O337.08 337.0 2-38 iPr H H H iPr C₂₃H₃₂N₂O 353.25 353.2 2-39 Cl H Cl Cl HC₁₇H₁₇Cl₃N₂O 371.04 371.0 2-40 Cl Cl H Cl Cl C₁₇H₁₆Cl₄N₂O 405.00 405.0

Example 3

Following the procedures described in the examples above, andsubstituting the appropriate starting materials and reagents, compounds3-1 to 3-11, having formula Ia, can also be prepared:

(Ia)

Ex. R² R³ R⁴ R⁵ R⁶ 3-1 F H F H F 3-2 F H H H F 3-3 F H Cl H F 3-4 F F HH F 3-5 F Cl H H F 3-6 F Me H H F 3-7 F H H F Cl 3-8 F H H H Cl 3-9 F HH H Br 3-10 F H Br H Cl 3-11 F H F H Cl

Example 4

Following the procedures described in the examples above, andsubstituting the appropriate starting materials and reagents, compounds4-1 to 4-26, having formula Ib, can also be prepared:

(Ib)

Ex. R^(1a) R^(1b) R^(1c) R^(1d) R² R³ R⁴ R⁵ R⁶ 4-1 F H H H F F H H F 4-2H F H H F H F H F 4-3 H F H H F H H H F 4-4 H F H H F H H H Cl 4-5 H H FH F H H H H 4-6 H H F H F H H H F 4-7 H H F H F H F H F 4-8 H H F H F FH H F 4-9 H H F H F H H F Cl 4-10 H H —CF₃ H F F H H F 4-11 H H —CF₃ H FH F H F 4-12 H H —CF₃ H F H H H Cl 4-13 H H H F F H H H H 4-14 H H H F FH H H Cl 4-15 H H H F F H H H F 4-16 H H H F F H F H F 4-17 H H H F F FH H F 4-18 H H H F F H H F Cl 4-19 H F F H F H H H F 4-20 H F F H F H FH F 4-21 H F F H F H H H Cl 4-22 H F H F F H H H F 4-23 H F H F F F H HF 4-24 H F H F F H F H F 4-25 H F H F F H H F Cl 4-26 H H F F F H F H F

Assay 1 hSERT, hNET, and hDAT Binding Assays

Membrane radioligand binding assays were used to measure inhibition oflabeled ligand (³H-citalopram or ³H-nisoxetine or ³H-WIN35428) bindingto membranes prepared from cells expressing the respective humanrecombinant transporter (hSERT or hNET or hDAT) in order to determinethe pK_(i) values of test compounds at the transporters.

Membrane Preparation From Cells Expressing hSERT, hNET, or hDAT

Recombinant human embryonic kidney (HEK-293) derived cell lines stablytransfected with hSERT or hNET, respectively, were grown in DMEM mediumsupplemented with 10% dialyzed FBS (for hSERT) or FBS (for hNET), 100μg/ml penicillin, 100 μg/ml streptomycin, 2 mM L-glutamine and 250 μg/mlof the aminoglycoside antibiotic G418, in a 5% CO₂ humidified incubatorat 37° C. When cultures reached 80% confluence, the cells were washedthoroughly in PBS (without Ca²⁺ and Mg²⁺) and lifted with 5 mM EDTA inPBS. Cells were pelleted by centrifugation, resuspended in lysis buffer(10 mM Tris-HCl, pH7.5 containing 1 mM EDTA), homogenized, pelleted bycentrifugation, then resuspended in 50 mM Tris-HCl, pH 7.5 and 10%sucrose at 4° C. Protein concentration of the membrane suspension wasdetermined using a Bio-Rad Bradford Protein Assay kit. Membranes weresnap frozen and stored at −80° C. Chinese hamster ovary membranesexpressing hDAT (CHO-DAT) were purchased from PerkinElmer and stored at−80° C.

Binding Assays

Binding assays were performed in a 96-well assay plate in a total volumeof 200 μL assay buffer (50 mM Tris-HCl, 120 mM NaCl, 5 mM KCl, pH 7.4)with 0.5, 1, and 3 μg membrane protein, for SERT, NET and DAT,respectively. Saturation binding studies, to determine radioligand K_(a)values for ³H-citalopram, ³H-nisoxetine, or ³H-WIN35428, respectivelywere conducted using 12 different radioligand concentrations rangingfrom 0.005-10 nM (³H-citalopram); 0.01-20 nM (³H-nisoxetine) and 0.2-50nM (³H-WIN35428). Inhibition assays for determination of pK_(i) valuesof test compounds were conducted with 1.0 nM ³H-citalopram, 1.0 nM³H-nisoxetine or 3.0 nM ³H-WIN35428, at 11 different concentrations oftest compound ranging from 10 μM to 100 μM.

Stock solutions (10 mM in DMSO) of test compound were prepared andserial dilutions made using Dilution Buffer (50 mM Tris-HCl, 120 mMNaCl, 5 mM KCl, pH 7.4, 0.1% BSA, 400 μM ascorbic acid). Non-specificradioligand binding was determined in the presence of 1 μM duloxetine, 1μM desipramine or 10 μM GBR12909 (each in Dilution Buffer) for thehSERT, hNET or hDAT assays, respectively.

Following a 60 minute incubation at 22° C. (or a period sufficient toreach equilibrium), the membranes were harvested by rapid filtrationover a 96-well UniFilter GF/B plate, pretreated with 0.3%polyethyleneimine, and washed 6 times with 300 μl wash buffer (50 mMTris-HCl, 0.9% NaCl, pH 7.5 at 4° C.). Plates were dried overnight atroom temperature, ˜45 μl of MicroScint™-20 (Perkin Elmer) added andbound radioactivity quantitated via liquid scintillation spectroscopy.Inhibition curves and saturation isotherms were analyzed using GraphPadPrism Software package (GraphPad Software, Inc., San Diego, Calif.).IC₅₀ values were generated from concentration response curves using theSigmoidal Dose Response (variable slope) algorithm in Prism GraphPad.K_(d) and B_(max) values for the radioligand were generated fromsaturation isotherms using the Saturation Binding Global Fit algorithmin Prism GraphPad. pK_(i) (negative decadic logarithm of K_(i)) valuesfor test compounds were calculated from the best-fit IC₅₀ values, andthe K_(d) value of the radioligand, using the Cheng-Prusoff equation(Cheng & Prusoff (1973) Biochem. Pharmacol. 22(23):3099-3108):K_(i)=IC₅₀/(1+[L]/K_(a)), where [L]=concentration radioligand.

Compounds of the invention were tested in this assay and found toexhibit SERT and NET pK_(i) values as follows:

SERT NET Ex. pK_(i) pK_(i) 1 ≧7.0 ≧7.0 2-1 ≧5.0 ≧7.0 2-2 ≧7.0 ≧5.0 2-3≧7.0 ≧7.0 2-4 ≧7.0 ≧5.0 2-5 ≧7.0 ≧5.0 2-6 ≧7.0 ≧5.0 2-7 ≧7.0 ≧5.0 2-8≧7.0 ≧5.0 2-9 ≧7.0 ≧5.0 2-10 ≧5.0 ≧7.0 2-11 ≧5.0 ≧5.0 2-12 ≧7.0 ≧7.02-13 ≧7.0 ≧7.0 2-14 ≧7.0 ≧5.0 2-15 ≧5.0 ≧5.0 2-16 ≧7.0 ≧5.0 2-17 ≧5.0≧5.0 2-18 ≧7.0 ≧5.0 2-19 ≧7.0 ≧5.0 2-20 ≧7.0 ≧5.0 2-21 ≧7.0 ≧7.0 2-22≧7.0 ≧5.0 2-23 ≧7.0 ≧5.0 2-24 ≧7.0 ≧5.0 2-25 ≧7.0 ≧7.0 2-26 ≧7.0 ≧5.02-27 ≧7.0 ≧5.0 2-28 ≧7.0 ≧5.0 2-29 ≧7.0 ≧5.0 2-30 ≧7.0 ≧7.0 2-31 ≧7.0≧5.0 2-32 ≧7.0 ≧5.0 2-33 ≧7.0 ≧7.0 2-34 ≧7.0 ≧7.0 2-35 ≧7.0 ≧7.0 2-36≧7.0 ≧5.0 2-37 ≧7.0 ≧5.0 2-38 ≧5.0 ≧5.0 2-39 ≧7.0 ≧7.0 2-40 ≧7.0 ≧5.0

Assay 2 hSERT, hNET, and hDAT Neurotransmitter Uptake Assays

Neurotransmitter uptake assays were used to measure inhibition of³H-serotonin (³H-5-HT), ³H-norepinephrine (³H-NE), and ³H-dopamine(³H-DA) uptake into cells expressing the respective transporter (hSERT,hNET or hDAT) in order to determine the pIC₅₀ values of test compoundsat the transporters.

³H-5-HT, ³H-NE, and ³H-DA Uptake Assays

HEK-293 derived cell lines stably-transfected with hSERT, hNET, or hDAT,respectively, were grown in DMEM medium supplemented with 10% dialyzedFBS (for hSERT) or FBS (for hNET and hDAT), 100 μg/ml penicillin, 100μg/ml streptomycin, 2 mM L-glutamine and 250 μg/ml of the aminoglycosideantibiotic G418 (for hSERT and hNET) or 800 ug/ml (for hDAT), in a 5%CO₂ humidified incubator at 37° C. When cultures reached 80% confluence,the cells were washed thoroughly in PBS (without Ca²⁺and Mg²⁺) andlifted with 5 mM EDTA in PBS. Cells were harvested by centrifugation at1100 rpm for 5 minutes, washed once by resuspension in PBS, thencentrifuged. The supernatant was discarded and the cell pelletresuspended, by gentle trituration, in room temperature Krebs-Ringerbicarbonate buffer containing HEPES (10 mM), CaCl₂ (2.2 mM), ascorbicacid (200 μM) and pargyline (200 μM), pH 7.4. The final concentration ofcells in the cell suspension was 7.5×10⁴ cells/ml, 1.25×10⁵ cells/ml,and 5.0×10⁴ cells/ml for SERT, NET, and DAT cell lines, respectively.

Neurotransmitter uptake assays were performed in a 96-well assay platein a total volume of 400 μL assay buffer (Krebs-Ringer bicarbonatebuffer containing HEPES (10 mM), CaCl₂ (2.2 mM), ascorbic acid (200 μM)and pargyline (200 μM), pH 7.4) with 1.5×10⁴ and 2.5×10⁴ cells, for SERTand NET, respectively. Inhibition assays for determination of pIC₅₀values of test compounds were conducted with 11 differentconcentrations, ranging from 10 μM to 100 μM. Stock solutions (10 mM inDMSO) of test compound were prepared and serial dilutions prepared using50 mM Tris-HCl, 120 mM NaCl, 5 mM KCl, pH 7.4, 0.1% BSA, 400 μM ascorbicacid. Test compounds were incubated for 30 minutes at 37° C. with therespective cells, prior to addition of radiolabeled neurotransmitter,³H-5-HT (20 nM final concentration), ³H-NE (50 nM final concentration),or ³H-DA (100 nM final concentration). Non-specific neurotransmitteruptake was determined in the presence of 2.5 μM duloxetine or 2.5 μMdesipramine (each in Dilution Buffer) for the hSERT, hNET, or hDATassays, respectively.

Following a 10 minute incubation, at 37° C., with radioligand, the cellswere harvested by rapid filtration over a 96-well UniFilter GF/B plate,pretreated with 1% BSA, and washed 6 times with 650 μl wash buffer (icecold PBS). Plates were dried overnight at 37° C., ˜45 μl ofMicroScint™-20 (Perkin Elmer) added and incorporated radioactivityquantitated via liquid scintillation spectroscopy. Inhibition curveswere analyzed using GraphPad Prism Software package (GraphPad Software,Inc., San Diego, Calif.). IC₅₀ values were generated from concentrationresponse curves using the Sigmoidal Dose Response (variable slope)algorithm in Prism GraphPad.

Compounds of the invention that were tested in this assay or in afluorescence-based assay as described in Tsuruda et al. (2010) Journalof Pharmacological and Toxicological Methods 61(2):192-204 (dataindicated by an asterisk in the table), were found to have serotonin andnorepinephrine reuptake inhibition pIC₅₀ values as follows:

Ex. SERT pIC₅₀ NET pIC₅₀ 2-12 ≧7.0* ≧7.0*

Assay 3 Ex Vivo SERT and NET Transporter Occupancy Studies

Ex vivo radioligand binding and neurotransmitter uptake assays are usedto determine the in vivo occupancy of SERT and NET, in selected brainregions, following in vivo administration (acute or chronic) of testcompounds. Following administration of test compound (by intravenous,intraperitoneal, oral, subcutaneous or other route) at the appropriatedose (0.0001 to 100 mg/kg), rats (≧n=4 per group) are euthanized atspecific time points (10 minutes to 48 hours) by decapitation and thebrain dissected on ice. Relevant brain regions are dissected, frozen andstored at −80° C. until use.

Ex Vivo SERT and NET Radioligand Binding Assays

For ex vivo radioligand binding assays, the initial rates of associationof SERT (³H-citalopram), and NET-(³H-nisoxetine) selective radioligandswith rat brain crude homogenates, prepared from vehicle and testcompound-treated animals, are monitored (see Hess et al. (2004) J.Pharmacol. Exp. Ther. 310(2):488-497). Crude brain tissue homogenatesare prepared by homogenizing frozen tissue pieces in 0.15 mL (per mg wetweight) of 50 mM Tris-HCl, 120 mM NaCl, 5 mM KCl, pH 7.4 buffer.Radioligand association assays are performed in a 96-well assay plate ina total volume of 200 μl assay buffer (50 mM Tris-HCl, 120 mM NaCl, 5 mMKCl, 0.025% BSA, pH 7.4) with 650 mg wet weight tissue (equivalent to 25μg protein). Homogenates are incubated for up to 5 minutes with³H-citalopram (3 nM) and ³H-nisoxetine (5 nM), respectively, prior totermination of the assay by rapid filtration over a 96-well UniFilterGF/B plate, pretreated with 0.3% polyethyleneimine. Filters then arewashed 6 times with 300 μl wash buffer (50 mM Tris-HCl, 0.9% NaCl, pH7.4 at 4° C.). Non-specific radioligand binding is determined in thepresence of 1 μM duloxetine, or 1 μM desipramine, for ³H-citalopram or³H-nisoxetine, respectively. The plates are dried overnight at roomtemperature, ˜45 μl of MicroScint™-20 (Perkin Elmer) is added, and boundradioactivity is quantitated via liquid scintillation spectroscopy. Theinitial rates of association of ³H-citalopram and ³H-nisoxetine aredetermined by linear regression using GraphPad Prism Software package(GraphPad Software, Inc., San Diego, Calif.). The average rate ofradioligand association to brain tissue homogenates from vehicle-treatedanimals is determined. The % occupancy of test compounds is thendetermined using the following equation:occupancy=100×(1−(initial rate association for test compound-treatedtissue/mean rate association for vehicle-treated tissue)ED₅₀ values are determined by plotting the log 10 of the dose of thetest compound against the % occupancy. ED₅₀ values are generated fromconcentration response curves using the Sigmoidal Dose Response(variable slope) algorithm in GraphPad Prism.

Ex Vivo SERT and NET Uptake Assays

Ex vivo neurotransmitter uptake assays, in which the uptake of ³H-5-HTor ³H-NE into rat brain crude homogenates, prepared from vehicle andtest compound-treated animals, are used to measure in vivo SERT and NETtransporter occupancy (see Wong et al. (1993) Neuropsychopharmacology8(1):23-33). Crude brain tissue homogenates are prepared by homogenizingfrozen tissue pieces in 0.5 mL (per mg wet weight) of 10 mM HEPES bufferpH 7.4, containing 0.32 M sucrose, 200 μM ascorbic acid and 200 μMpargyline, at 22° C. Neurotransmitter uptake assays are performed in a96-well Axygen plate in a total volume of 350 μl assay buffer(Krebs-Ringer bicarbonate buffer with 10 mM HEPES, 2.2 mM CaCl₂, 200 μMascorbic acid and 200 μM pargyline, pH 7.4) with 50 μg protein.Homogenates are incubated for 5 minutes at 37° C. with ³H-5-HT (20 nM)and ³H-NE (50 nM), respectively, prior to termination of the assay byrapid filtration over a 96-well UniFilter GF/B plate, pretreated with 1%BSA. Plates are washed 6 times with 650 μl wash buffer (ice cold PBS)and dried overnight at 37° C., prior to addition of ˜45 μl ofMicroScint™-20 (Perkin Elmer). Incorporated radioactivity is quantitatedvia liquid scintillation spectroscopy. Non-specific neurotransmitteruptake is determined in parallel assays in which tissue homogenates areincubated with ³H-5-HT (20 nM) or ³H-NE (50 nM) for 5 minutes at 4° C.

Assay 4 Other Assays

Other assays that are used to evaluate the pharmacological properties oftest compounds include, but are not limited to, cold ligand bindingkinetics assays (Motulsky and Mahan (1984) Molecular Pharmacol.25(1):1-9) with membranes prepared from cells expressing hSERT or hNET;conventional membrane radioligand binding assays using radiolabeled, forexample, tritiated, test compound; radioligand binding assays usingnative tissue from, for example rodent or human brain; neurotransmitteruptake assays using human or rodent platelets; neurotransmitter uptakeassays using crude, or pure, synaptosome preparations from rodent brain.

Assay 5 Formalin Paw Test

Compounds are assessed for their ability to inhibit the behavioralresponse evoked by a 50 μl injection of formalin (5%). A metal band isaffixed to the left hind paw of male Sprague-Dawley rats (200-250 g) andeach rat is conditioned to the band for 60 minutes within a plasticcylinder (15 cm diameter). Compounds are prepared in pharmaceuticallyacceptable vehicles and administered systemically (i.p., p.o.) atpre-designated times before formalin challenge. Spontaneous nociceptivebehaviors consisting of flinching of the injected (banded) hind paw arecounted continuously for 60 minutes using an automated nociceptionanalyzer (UCSD Anesthesiology Research, San Diego, Calif.).Antinociceptive properties of test articles are determined by comparingthe number of flinches in the vehicle and compound-treated rats (Yakshet al., “An automated flinch detecting system for use in the formalinnociceptive bioassay” (2001) J. Appl. Physiol. 90(6):2386-2402).

Assay 6 Spinal Nerve Ligation Model

Compounds are assessed for their ability to reverse tactile allodynia(increased sensitivity to an innocuous mechanical stimulus) induced bynerve injury. Male Sprague-Dawley rats are surgically prepared asdescribed in Kim and Chung “An experimental model for peripheralneuropathy produced by segmental spinal nerve ligation in the rat”(1992) Pain 50(3):355-363. Mechanical sensitivity is determined as the50% withdrawal response to innocuous mechanical stimuli (Chaplan et al.,“Quantitative assessment of tactile allodynia in the rat paw” (1994) J.Neurosci. Methods 53(1):55-63) before and after nerve injury. One tofour weeks post-surgery, compounds are prepared in pharmaceuticallyacceptable vehicles and administered systemically (i.p., p.o.). Thedegree of nerve injury-induced mechanical sensitivity before and aftertreatment serves as an index of the compounds' antinociceptiveproperties.

While the present invention has been described with reference tospecific aspects or embodiments thereof, it will be understood by thoseof ordinary skilled in the art that various changes can be made orequivalents can be substituted without departing from the true spiritand scope of the invention. Additionally, to the extent permitted byapplicable patent statues and regulations, all publications, patents andpatent applications cited herein are hereby incorporated by reference intheir entirety to the same extent as if each document had beenindividually incorporated by reference herein.

What is claimed is:
 1. A compound of formula I:

where: a is 0, 1, or 2; each R¹ is independently halo ortrifluoromethyl; R² is hydrogen, halo, —C₁₋₆alkyl, —O—C₁₋₆alkyl,—C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl, or —CN; R³ is hydrogen,halo, —C₁₋₆alkyl, or —O—C₁₋₆alkyl; R⁴ is hydrogen, halo, or —C₁₋₆alkyl;R⁵ is hydrogen or halo; and R⁶ is hydrogen, halo, or —C₁₋₆alkyl; or apharmaceutically acceptable salt thereof; with the proviso that thecompound of formula I is not 1-(2-phenoxymethylphenyl)piperazine,1-[2-(3-fluorophenoxymethyl)phenyl]piperazine, or1-[2-(2,3-difluorophenoxymethyl)phenyl]-piperazine.
 2. The compound ofclaim 1, where a is
 0. 3. A pharmaceutical composition comprising acompound of claim 1 and a pharmaceutically acceptable carrier.
 4. Thepharmaceutical composition of claim 3 further comprising a therapeuticagent selected from anti-Alzheimer's agents, anticonvulsants,antidepressants, anti-Parkinson's agents, dual serotonin-norepinephrinereuptake inhibitors, non-steroidal anti-inflammatory agents,norepinephrine reuptake inhibitors, opioid agonists, opioid antagonists,selective serotonin reuptake inhibitors, sodium channel blockers,sympatholytics, and combinations thereof.
 5. A compound of formula IV,useful in the synthesis of a compound of formula I:

where: a is 0, 1, or 2; each R¹ is independently halo ortrifluoromethyl; R²⁻⁶ are independently hydrogen, halo, —C₁₋₆ alkyl,—CF₃ , —C₂₋₆ alkynyl, —O—C₁₋₆ alkyl, —OCF₃, —C₁₋₄ alkylene-O—C₁₋₄ alkyl,—C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl, —C₀₋₆alkylene-OH, —CN,—C₀₋₂alkylene-COOH, —CHO, —C(O)—C₁₋₆alkyl, —C(O)O—C₁₋₄alkyl, —CH₂SH,—S—C₁₋₆alkyl, —C₁₋₄alkylene—S—C₁₋₄alkyl, —SO₂—C₁₋₆alkyl, —SO₂NRR,—NHSO₂R, —C₀₋₁alkylene—NRR, —NHC(O)—C₁₋₆alkyl, —C(O)NRR, or —NO₂; andeach R is independently hydrogen or —C₁₋₄alkyl; with the proviso thatthe compound of formula I is not 1-(2-phenoxymethylphenyl)piperazine,1-[2-(3-fluorophenoxymethyl)phenyl]piperazine, or1-[2-(2,3-difluorophenoxymethyl)phenyl]-piperazine;

where P is an amino-protecting group selected from t-butoxycarbonyl,trityl, benzyloxycarbonyl, 9-fluorenylmethoxycarbonyl, formyl, andbenzyl.
 6. The compound of claim 5, where a is 0; and R²⁻⁶ areindependently hydrogen, halo, —C₁₋₆alkyl, —O—C₁₋₆alkyl,—C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl, or —CN.
 7. A method ofpreparing a compound of formula I:

where: a is 0, 1, or 2; each R¹ is independently halo ortrifluoromethyl; R²⁻⁶ are independently hydrogen, halo, —C₁₋₆alkyl,—CF₃, —C₂₋₆alkynyl, —O—C₁₋₆alkyl, —OCF₃, —C₁₋₄alkylene-O—C₁₋₄alkyl,—C₀₋₁alkylene-phenyl, —O—C₀₋₃alkylene-phenyl, —₀₋₆alkylene-OH,—CN,—₀₋₂alkylene-COOH, —CHO, —C(O)—C₁₋₆alkyl, —C(O)O—C₁₋₄alkyl, —CH₂SH,—S—C₁₋₆alkyl, —C₁₋₄alkylene—S—C₁₋₄alkyl, —SO₂—C₁₋₆alkyl, —SO₂NRR,—NHSO₂R, —C₀₋₁alkylene-NRR, —NHC(O)—C₁₋₆alkyl, —C(O)NRR, or —NO₂; andeach R is independently hydrogen or —C₁₋₄alkyl; with the proviso thatthe compound of formula I is not 1-(2-phenoxymethylphenyl)piperazine,1-[2-(3-fluorophenoxymethyl)phenyl]piperazine, or1-[2-(2,3-difluorophenoxymethyl)phenyl]-piperazine; the processcomprising deprotecting a compound of formula IV:

or a salt thereof, to provide a compound of formula I; where P is anamino-protecting group selected from t-butoxycarbonyl, trityl,benzyloxycarbonyl, 9-fluorenylmethoxycarbonyl, formyl, and benzyl. 8.The method of claim 7, where a is 0; and R²⁻⁶ are independentlyhydrogen, halo, —C₁₋₆alkyl, —O—C₁₋₆alkyl, —C₀₋₁alkylene-phenyl,—O—C₀₋₃alkylene-phenyl, or —CN.